Báo cáo khoa học: Serine-arginine protein kinases: a small protein kinase family with a large cellular presence potx

REVIEW ARTICLE
Serine-arginine protein kinases: a small protein kinase
family with a large cellular presence
Thomas Giannakouros
1
, Eleni Nikolakaki
1
, Ilias Mylonis
2
and Eleni Georgatsou
2
1 Laboratory of Biochemistry, Department of Chemistry, Aristotle University of Thessaloniki, Greece
2 Laboratory of Biochemistry, Department of Medicine, School of Health Sciences, University of Thessaly, Larissa, Greece
History of the discovery of the
serine-arginine protein kinase
(SPRK) family
The first serine-arginine (SR) protein kinase to be puri-
fied and characterized was named SRPK1, for SR-pro-
tein-specific kinase 1 [1,2]. It was isolated during a
search for the activity that phosphorylates SR splicing
factors (also named SR proteins) during mitosis.
SRPK1 was shown to phosphorylate SR proteins in a
cell-cycle regulated manner, to affect SR protein locali-
zation and to inhibit splicing when added in large
quantities to a cell-free splicing assay [1,2]. The
SRPK1 cDNA was cloned, revealing that the Schizo-
saccharomyces pombe SRPK1 orthologue, Dsk1, had
already been cloned and partially characterized as a
kinase with cell cycle-dependent phosphorylation and
subcellular localization [3]. The SRPK1 and Dsk1
nucleotide sequencing identified a domain interrupting

the kinase catalytic site into two structural entities,
Keywords
LBR; metabolic signalling; nuclear envelope;
p53; PGC-1; protamine; spermatogenesis;
splicing; SR protein; SRPK
Correspondence
E. Georgatsou, Laboratory of Biochemistry,
Department of Medicine, School of Health
Sciences, University of Thessaly, Biopolis,
41110 Larissa, Greece
Fax: +30 2410 685545
Tel: +30 2410 685581
E-mail:
(Received 7 July 2010, accepted 26 October
2010)
doi:10.1111/j.1742-4658.2010.07987.x
Serine-arginine protein kinases (SPRKs) constitute a relatively novel
subfamily of serine-threonine kinases that specifically phosphorylate serine
residues residing in serine-arginine ⁄ arginine-serine dipeptide motifs. Fifteen
years of research subsequent to the purification and cloning of human
SRPK1 as a SR splicing factor-phosphorylating protein have lead to the
accumulation of information on the function and regulation of the different
members of this family, as well as on the genomic organization of SRPK
genes in several organisms. Originally considered to be devoted to constitu-
tive and alternative mRNA splicing, SRPKs are now known to expand
their influence to additional steps of mRNA maturation, as well as to other
cellular activities, such as chromatin reorganization in somatic and sperm
cells, cell cycle and p53 regulation, and metabolic signalling. Similarly,
SRPKs were considered to be constitutively active kinases, although several
modes of regulation of their function have been demonstrated, implying an

elaborate cellular control of their activity. Finally, SRPK gene sequence
information from bioinformatics data reveals that SRPK gene homologs
exist either in single or multiple copies in every single eukaryotic organism
tested, emphasizing the importance of SRPK protein function for cellular
life.
Abbreviations
CDK, cyclin dependent kinase; Clk, CDK-like kinase; CK2, casein kinase 2; FOXO1, forkhead box protein O1; HBV, hepatitis B virus;
HP1, heterochromatin protein 1; Hsp, heat shock protein; LBR, lamin B receptor; NRF-1, nuclear respiratory factor-1; PGC-1, peroxisome
proliferator activated receptor c coactivator-1; RS, arginine-serine; SAFB, scaffold attachment factor B; SR, serine-arginine;
SRPK, serine-arginine protein kinase.
570 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
hence called ‘the spacer domain’, which is characteris-
tic of the SR protein kinase family [3,4]. Subsequently,
Dsk1 was also shown to be a SR protein kinase phos-
phorylating and regulating the function of SR proteins
[5–7].
In 1998, the cloning of SRPK2 was reported almost
simultaneously in mouse (together with mSRPK1) [8]
and man [9]. SRPK2 was found to be a SR-specific
protein kinase highly homologous to SRPK1. It is
structurally differentiated from SRPK1 by a proline-
rich tract at its N-terminus and an acidic region in its
spacer domain. However, none of these elements had
been related to a particular SRPK2-specific function
until recently, when a study showed that sequences
residing in the acidic domain of SRPK2 specifically
interact with the pro-apoptotic arginine-serine (RS)
domain-containing protein acinus [10]. Moreover, the
mRNA of SRPK2 was shown to have a different
and more limited tissue distribution than SRPK1

mRNA [9].
The cloning and characterization of the budding
yeast Saccharomyces cerevisiae SR protein kinase Sky1
in 1999 came not only unexpectedly, but also as a
challenge because the prevalence of uninterrupted
genes and the lack of alternative splicing in this organ-
ism would not, at that time, account for a SR protein-
specific kinase [11]. Sky1 was indeed shown to have
the structural and functional characteristics of a SR
protein kinase because it could not only phosphorylate
mammalian SR splicing factors in vivo [12], but also
native RS domain-containing S. cerevisiae proteins,
such as Npl3p, which is involved in mRNA export
[13]. This observation led to the discovery of the
involvement of SRPKs in the regulation of additional
steps of mRNA maturation and added to the current
image of coupled transcript processing from the tran-
scription to translation steps.
The cloning of SPK-1, the unique homolog of
SRPK1 in Caenorhabditis elegans, revealed an essential
function of the kinase in the germline development
and embryogenesis of this organism [14]. The underly-
ing mechanism for this function has not yet been eluci-
dated, although the finding that human SRPK1 is
highly expressed in testis and phosphorylates prot-
amine 1, a highly basic protein replacing histones dur-
ing spermiogenesis, could be related with the
observations in C. elegans [15].
SRPK1a, a product of the SRPK1 gene produced by
alternative splicing, that retains an additional domain

corresponding to an intron at its N-terminal region,
was reported in 2001 [16]. Interestingly, this domain is
rich in proline residues reminiscent of the proline-rich
SRPK2-specific track. Additionally, the SRPK1a
N-terminus was found to interact with the nuclear
matrix protein scaffold attachment factor (SAFB) B1,
and it was subsequently shown that SAFB proteins are
inhibitors of SRPK1 and SRPK1a activity, function-
ally differentiating between the two kinases and further
implicating SRPKs in subnuclear organization and
chromatin regulation [17].
Mouse SRPK3 was discovered in 2005, having been
identified in a screen for target genes of the transcrip-
tion factor myocyte enhancer factor 2 [18]. SRPK3 is
expressed in a tissue-specific fashion in the heart and
skeletal muscle and is required for normal muscle
growth and homeostasis because Srpk3-null mice suffer
from centronuclear myopathy [18]. It has not been
confirmed, however, whether SR kinase activity is
required for these phenotypes and, if so, what sub-
strates are affected. The existence of the orthologue of
mSRPK3 in humans has been postulated in an analysis
of human chromosomal DNA methylation, although
no studies are available for its expression or function.
However, the cDNA of the porcine SRPK3 has been
cloned and shown to have a very limited and tissue
specific expression in muscular tissue [19].
Although Drosophila harbors several SRPK homo-
logs, only two very recent studies refer to Srpk79D (as
named by both groups), which is considered to be a

product of the CG11489 gene in the Drosophila
genome [20,21]. It is interesting that Srpk79D displays
tissue specific expression in neuronal tissue and is
implicated in the development and growth of synaptic
connections throughout the nervous system.
Finally, some SR protein kinases of lower organisms
have also been cloned, adding to the picture of SRPK
function and importance. TcSRPK, the SR protein
kinase of a protozoan, the parasite Trypanosoma cruzi,
which displays trans- and cis-splicing and was cloned
and characterized in 2003, functions as a bona fide SR
protein kinase, indicating that the general control of
eukaryotic mRNA processing evolved early during
evolution [22]. More recently, PSRPK, the SR protein
kinase of Physarum polycephalum, a slime mold, has
been cloned and characterized, especially with respect
to its subcellular localization properties [23].
Evolution of the SRPK gene family
A simple search for genes (using the keyword ‘SRPK*’
at ) returns approximately
90 hits for SRPK genes or putative SRPK genes in dif-
ferent eukaryotic organisms. Some of these sequence
entries have not yet been subjected to a final NCBI
review and overlaps may exist between them that
should be thoroughly examined. In our preliminary
T. Giannakouros et al. Serine-arginine protein kinases
FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS 571
research, however, we have made some interesting
observations that we note below to emphasize the sig-
nificance of evolutionary-oriented studies on the

sequences of the SRPK gene products.
The first intriguing observation is that the SRPK gene
copy number of an organism does not appear to directly
relate to its evolutionary scale. For example, there
exist fungi with one, two or even up to nine SRPK
genes [S. cerevisiae and S. pombe with one gene (Sky1
and Dsk1, respectively); Candida albicans with two
(QSAA48 and QS9Q27); Aspergilus niger with nine
(A2QAE4, A2QB94, A2QC46, A5AB23, A2QWQ2,
A2QX01, A2QX98, A2R2M0 and A2RSV1)]; plants
with three genes (Ricinus communis; B9SRL4, B9S6V7
and B9SNS8); and insects with two (Culex quinquefasci-
atus; BOWGI3 and BOWRV4) or three genes [Drosoph-
ila melanogaster; CG8174, CG8565 and CG11489
(CG9085)], whereas mammals (rat, mouse, human, etc.)
have three genes. Additionally, as we have experienced
from our own research and as m entioned in the two s tudies
concerning Srpk79D in Drosophila melanogaster [20,21],
there is no prominent one-to-one correspondence
between the sequences of SRPK genes of evolutionary
remote species. The emerging image is reminiscent of
independent SRPK gene duplications that have taken
place at several time points during evolution in different
species. Accordingly, it is suggested that the SRPK
genes are subjected to an evolutionary drive that
demands multiple SRPK gene copies in almost each new
emerging species. One may observe evidence of the
errors of the evolutionary ‘trial and error’ process oper-
ating through new SRPK genes: pseudogenes exist for
both SRPK1 and SRPK2 in the human genome and

also for SRPK1 in the mouse [24]. Other loci identified
by sequencing might also correspond to pseudogenes.
The second observation concerns the ‘spacer region’
of the SRPK proteins. This sequence is SRPK family-
specific (as a serine ⁄ threonine kinase subfamily) and
each family member harbors its own unique spacer. It
should be noted that the different ‘spacer regions’, in
addition to being very different with respect to their
primary sequence, are very diverse in length, and pos-
sibly function too, as indicated by the data presented
further below. Consequently, it is not unexpected that,
for all the SRPK sequences we randomly examined
from different kingdoms, the whole spacer sequence
resides on a separate exon, suggesting that this domain
may have evolved independently.
Another domain of interest that remains relatively
unexamined from an evolutionary point of view is the
N-terminal domain of the kinases. It is highly specific
between the SRPK family members, and few functions
have been attributed to it. It may be important to note
that it is this particular region of the mRNA that fre-
quently swaps and alternates in the splicing phenom-
ena that are beginning to be revealed in SRPK
transcripts [16,20,21].
Finally, it is interesting to note that SRPK2 contains
a minor class of introns [25]. Because the minor class
of introns is often associated with many important
genes that are evolutionarily conserved, it is likely that
SRPK2 is evolving and regulated by a distinct mecha-
nism from SRPK1.

Function of the SRPKs
As already noted, SRPKs phosphorylate their
substrates at serine residues located in regions rich in
arginine ⁄ serine dipeptides, known as RS domains. The
definition of a ‘typical’ RS domain is somewhat arbi-
trary and SRPKs have been shown to be able to phos-
phorylate scattered RS dipeptides if they conform to
certain limitations [26–29]. The specificity of these
enzymes is remarkable because mutations of Ser to
Thr or Arg to Lys in the RS domain completely abro-
gate phosphorylation [2,26].
In the list of the RS domain-containing proteins, the
SR proteins prevail, either as the originally identified
‘classical’ SR proteins invariably containing an RNA
recognition motif or as ‘SR-like’ or ‘SR-related’ pro-
teins also containing RNA binding domains (RNA
recognition motif or other). Most of the SR splicing
factors have been experimentally shown to be SRPK
substrates in vitro and in vivo and it is to be expected
that every SR protein could potentially be a SRPK
substrate under particular cellular conditions. Yet a
recent study suggests that the human genome encodes
for more than 100 RS domain-containing proteins [30],
indicating that SRPKs may regulate diverse cellular
functions through phosphorylation of many of these
potential substrates. Below, we review the SRPK
impact on mRNA maturation and discuss the regula-
tory paradigms that have been characterized to a
reasonable extent, including the replacement of hist-
ones by the arginine-rich protamines during spermio-

genesis, the role of SPRKs in cell cycle progression
and chromatin reorganization, and the function of
SRPKs in the regulation of peroxisome proliferator
activated receptor c coactivator (PGC)-1a in metabolic
signaling.
SRPKs and mRNA maturation
The involvement of SRPKs in the regulation of
mRNA splicing was expected because SRPK1 was iso-
lated as a SR splicing factor-phosphorylating kinase
Serine-arginine protein kinases T. Giannakouros et al.
572 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
and the phosphorylation of SR proteins had been
shown to be a prerequisite for spliceosome assembly
and splicing in general [31–33]. However, the exact
contribution of the SRPKs in the different steps of
mRNA maturation is not completely clarified (even up
to this date) for several reasons. First, the subcellular
localization of SRPKs is cytoplasmic as well as
nuclear, implying a more complex function for these
kinases than the phosphorylation of only cytoplasmic-
or only nuclear-localized SR splicing factors. SRPKs
undoubtedly phosphorylate both, although under con-
ditions that are strictly controlled. Second, SR protein
phosphorylation in the nucleus also takes place as a
result of other families of kinases. The cyclin depen-
dent kinase (CDK)-like kinases (Clk) also phosphory-
late RS domains but have a much broader specificity
[26,34]. Topoisomerase I has also been shown to
phosphorylate SR proteins [35] but its role in SR
protein function remains unexplored and, finally, and

also relatively recently, Akt kinases have been shown
to affect splicing by targeting RS domains [36]. Third,
the specific functions of the various SRPKs discov-
ered in different organisms are just beginning to be
addressed.
As already mentioned, concomitant with its purifica-
tion, SRPK1 was shown to inhibit splicing in vitro
when present in large quantities and to disassemble
nuclear speckles when added in permeabilized cells [1].
This and other in vitro experiments have implicated
SRPKs in the phosphorylation of SR splicing factors
and the regulation of splicing [2,9,37], although the
first study to definitively attribute a role of a SRPK on
SR protein function in vivo was carried out by Yeakley
et al. [12], which showed that when the unique SRPK
of S. cerevisiae (Sky1) is deleted, the interaction of SR
proteins is prevented, and they are incapable of trans-
locating into the nucleus. Importantly, that study,
which used mammalian splicing factors, showed for
the first time that SRPK-mediated phosphorylation
plays an important role in SR protein nuclear import
and that not all SR splicing factors are affected identi-
cally. Sequential studies with Sky1 and its SR-like pro-
tein substrate Nlp3p (which transports mRNAs out of
the nucleus) have shown that Nlp3p needs to be phos-
phorylated to release the mRNA and be re-imported
into the nucleus [13,38]. In humans, shuttling splicing
factors such as SF2 ⁄ ASF are phosphorylated in the
cytoplasm by SRPK1 (Fig. 1) and are subsequently
Fig. 1. SRPK regulation and function in mRNA maturation. During interphase, SRPKs are sequestered in the cytoplasm via their spacer

domain and anchoring to various protein complexes, where they phosphorylate SR proteins and facilitate their nuclear import. After stress-
induced, cell cycle-dependent or other signalling, SRPKs translocate to the cell nucleus where, along with other SR protein kinases (Clks),
they further modify their substrates found in nuclear speckles. SRPK-mediated phosphorylation influences the dissociation of SR splicing fac-
tors from speckles, spliceosome assembly and splice site selection. Dephosphorylation of SR splicing factors is required for splicing activity
and their export to the cytoplasm. In the nucleus, SRPKs can interact with nuclear matrix proteins such as SAFB. Their export is the result
of an as yet unidentified mechanism. Black arrows indicate molecule reactions or movements. Dashed lines indicate hypothetical molecule
reactions or movements.
T. Giannakouros et al. Serine-arginine protein kinases
FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS 573
transported to the nucleus by transportin-SR2, which
specifically interacts with phosphorylated RS domains
[39–41].
In the nucleus, when not actively involved in tran-
script processing, SR proteins reside in nuclear speckles
from which they are released when subjected to a new
round of phosphorylation. Although SRPKs are able
to phosphorylate SR proteins residing in nuclear speck-
les, it is difficult to determine the exact roles of the
different kinases involved in phosphorylation of SR
splicing factors in the nucleus (Fig. 1). The importance
of the function of SRPKs for the splicing reaction
in vivo first became apparent from the genetic approach
of Dagher & Fu [42] in budding yeast, where it was
shown that Sky1 interacts with proteins that affect
3¢ splice site selection. Additionally, in co-transfection
experiments, Nikolakaki et al. [16] showed that both
SRPK1 and SRPK1a are capable of altering the splic-
ing of a tau minigene in a dose-dependent manner.
Moreover, it is interesting that, in humans, SRPK1 has
been found to be associated with the U1-snRNP, which

is involved in 5¢ splice selection [43], whereas SRPK2
was shown to be required for the formation of the
U4 ⁄ U6-U5 tri-snRNP, which is involved in 3¢ splice site
selection [44]. Similarly, using siRNA, Hayes et al. [45]
have confirmed the role of SRPK1 in phosphorylating
SR proteins in vivo and have connected the endogenous
down-regulation of SRPK1 expression with alternative
splicing of a particular transcript. Finally, Zhong et al.
[46] clearly showed that when SR protein kinases enter
the nucleus (in this case under a stress signal), phos-
phorylation of SR proteins is increased, verifying the
nuclear action of SRPKs on SR splicing factors.
Accordingly, a study by Jiang et al. [47] showed that
when SRPK2 is phosphorylated by Akt in neuronal
cells, it enters the nucleus and is able to phosphorylate
the nonshuttling SR splicing factor SC35.
As previously noted, SR proteins are implicated in a
much broader spectrum of activities that accompany
the life of an mRNA, in addition to the splicing pro-
cess. To function in mRNA export, SR proteins need
to be underphosphorylated (Fig. 1). On the other
hand, SF2 ⁄ ASF does not have to leave the nucleus to
exert its positive effect on mRNA nonsense mediated
decay [48]. An intact RS domain is required for this
particular function, yet the impact of its phosphoryla-
tion state has not been addressed. In addition,
SF2 ⁄ ASF has been recently shown to be a transla-
tional activator of capped mRNAs in the cytoplasm
[49]; however, no report on its state of phosphoryla-
tion was included in that study. The participation of

the SRPKs in these SR protein-dependent functions
would be an interesting subject for future studies. In
this respect, it should be noted that, in yeast, where
the SR-like protein Npl3 was found to also affect
translation (albeit by a different mechanism than
SF2 ⁄ ASF in humans), this activity was shown to be
Sky1-independent [50].
The key role played by SRPKs in mRNA processing
is particularly apparent in studies on pathological con-
ditions, such as viral infection and tumor development.
The herpes simplex virus-1 protein ICP27 interacts
with SRPK1, relocalizing it to the nucleus and affect-
ing its function, resulting in lower total host splicing
activity, and thus favoring the exit from the nucleus of
the intronless viral mRNAs [51]. The E1^E4 protein of
human papilloma virus 1 interacts with SRPK1 and
can function as a substrate for the kinase. The in vivo
effects of this interaction are not known, however, nor
is it known whether these putative effects would be
exerted via the splicing machinery [52]. A third virus
found to be directly involved in SRPK function is hep-
atitis B virus (HBV). In HBV-infected cells, before the
encapsidation of the virus genetic material, the unique
viral core protein needs to be phosphorylated by a
host kinase. Although there are conflicting results as to
whether the kinases responsible for this phosphoryla-
tion are SRPK1 and 2, there is agreement on the fact
that the viral protein interacts with SRPK1 and that
this interaction affects the HBV cell cycle [53,54]. This
as well as other evidence suggests that SRPKs may be

potential pharmaceutical targets for the control of viral
infection. Hence, a small molecule, isonicotinamide
compound, which is a relatively selective inhibitor of
SRPK1 and 2 (SPRIN340), was found to impair Sind-
bis virus propagation in cultured cells, although it is
only variably effective on HIV-1 propagation [55].
Interest in SRPKs as pharmaceutical targets also
emerged from the observation that SRPKs show
increased expression in tumors of pancreas, breast and
colon [45,56], as well as in acute T-cell leukemia
induced by human T-cell leukemia virus-1 [57].
Accordingly, cell lines derived from pancreatic, breast
and colonic tumors, when disrupted for the SRPK1
gene, display diminished cell proliferation, increased
apoptotic potential and augmented sensitivity to the
common chemotherapeutics gemcitabine and cisplatine.
Evidence has been provided that the results observed
are effected through the splicing machinery [45]. An
inverse correlation has been documented, however,
between the expression of SRPK1 and cisplatin sensi-
tivity in yeast and in cells of germline origin, where
down-regulation of SRPK1 confers resistance to cis-
platin [58,59]. These tissue-specific findings again point
out the intricate and fine-tuned cellular networks regu-
lated by SRPK activity.
Serine-arginine protein kinases T. Giannakouros et al.
574 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
SRPKs and spermiogenesis
SRPK1, SRPK1a and SRPK2 are predominantly
expressed in testis [9,15,16]. Because of the numerous

maturation stages that germ cells undergo, novel gene
regulation strategies have been developed that provide
for flexible gene expression and protein function and,
among them, alternative splicing is particularly preva-
lent. The overexpression of SRPKs in testis may there-
fore suggest a contributory role for these kinases in
generating altered splice patterns across the develop-
mental program of germ cells. Yet the levels of SR
proteins are not elevated in testis compared to other
tissues [60,61], implying that the high levels of SRPKs
are probably not just a result of changes in the splicing
machinery.
The finding that P1 protamine from various organ-
isms satisfy the substrate specificity requirements of
SRPK1, coupled with the fact that most, if not all, of
these proteins are known to be phosphoproteins
[62,63], made them attractive candidate substrates of
SRPK1. Consistent with this hypothesis, Ser10 and
Ser8 (
7
RSQSRSR
13
) were identified as the in vivo
phosphorylation sites of mono- and di-phosphorylated
human P1 protamine [62]. Indeed, SRPK1 was found
to phosphorylate human P1 protamine efficiently [15].
Protamines are highly basic, arginine-rich, low-molecu-
lar weight proteins that replace histones during the
development of spermatids into spermatozoa, a process
termed spermiogenesis [63]. As a result of this

exchange, the nucleosomal-type chromatin is trans-
formed into a smooth fiber and compacted into a
volume approximately 5% of that of a somatic cell
nucleus [63,64]. P1 protamine is the main member of
the family and is conserved in all vertebrates, whereas
P2 protamine has been described only in some species,
including man, stallion, hamster and mouse [63].
The deposition of protamines on sperm chromatin
and the subsequent chromatin condensation are largely
controlled by phosphorylation-dephosphorylation
events. Protamines are highly phosphorylated shortly
after their synthesis and before binding to DNA [65].
Phosphorylation of P2 protamine has been shown to
be essential because deletion of the calmodulin-depen-
dent protein kinase Camk4, which phosphorylates P2
protamine, impairs the deposition of P2 protamine on
sperm chromatin, resulting in defective spermiogenesis
and male sterility [66]. Phosphorylation of P1 prot-
amine by SRPK1 is required for the temporal associa-
tion of P1 protamine with lamin B receptor (LBR), an
inner nuclear membrane protein that also possesses a
stretch of RS dipeptides at its nucleoplasmic NH
2
-
terminal domain [67]. It is well known that RS
domains mediate protein–protein interactions in a
phosphorylation-dependent manner [68], assuming that
only one of the two RS domains is phosphorylated.
Phosphorylation of the P1 protamine molecules in the
cytoplasm on their way to the nucleus together with a

lack of LBR phosphorylation is consistent with the
observed predominant cytoplasmic localization of
SRPK1 and the minimal RS kinase activity detected in
the nucleus of germ cells [4,15].
The association of P1 protamine with the nuclear
envelope probably represents an important intermedi-
ate step before its deposition on sperm chromatin. In
this respect, Biggiogera et al. [69] reported that prota-
mines initially appear at the nuclear periphery, imply-
ing that the nuclear envelope might play a role in the
replacement of transition proteins by protamines dur-
ing spermiogenesis. The detachment of P1 protamine
from the nuclear envelope and its binding to DNA are
probably achieved through its dephosphorylation
(Fig. 2). Consistent with this hypothesis, protamines
were found mainly dephosphorylated in mature sperm
chromatin [62,63]. One possibility is that the nuclear
envelope functions as a ‘working platform’ where addi-
tional modifications (e.g. methylation) of P1 protamine
take place. These modifications may not only increase
the affinity of P1 protamine for sperm DNA, but also
may recruit specific molecules, such as heterochromatin
protein 1 (HP1), which were shown to be coupled to
chromatin condensation and transcriptional silencing
[64,70].
A central question concerning P1 protamine is how
its transportation into the nucleus is accomplished.
Conceivably, this may be mediated through an active
transport mechanism, similar to histone H1 and transi-
tion protein 2, for which importin 5 and importin 4,

respectively, are known to be responsible for their
translocation into the nucleus [71,72]. Consistent with
this hypothesis, it has been suggested that phosphory-
lation of the RS domain of the splicing factor
ASF ⁄ SF2 by SRPK1 results in a conformational
change that facilitates its interaction with the nuclear
transport receptor transportin-SR2 (an importin-b
family protein), thereby mediating the shuttling of this
SR protein into the nucleus through the nuclear pore
complex [41]. In such a case, phosphorylation of P1
protamine by cytoplasmic SRPK1 may also promote
its interaction with an as yet unknown importin family
member, thereby facilitating its translocation into the
nucleus. The release of P1 protamine from importin
may be mediated through its binding to LBR at the
nuclear periphery.
Finally, SRPKs may have additional roles in sper-
matogenesis that need to be further characterized. For
T. Giannakouros et al. Serine-arginine protein kinases
FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS 575
example, SRPK1 was reported to mediate the uptake
of polyamines through an as yet unidentified signaling
pathway [73].
SRPKs, cell cycle progression and chromatin
reorganization
SRPKs have been characterized as cell cycle regulated
kinases [1,3]. This characterization was mainly based
on the finding that SRPK1, as well as its fission yeast
homolog, Dsk1, can translocate into the nucleus at the
end of the G2 phase [3,4]. In addition, SRPK1 activity,

when assayed using SC35 or ASF ⁄ SF2 as substrate,
has been reported to be approximately five-fold higher
in extracts from metaphase compared to interphase
cells [1]. The break-up of the speckled pattern and the
redistribution of splicing factors throughout the cyto-
plasm were initially considered as the main mitotic
functions of SRPK1 [1]. In this review, we discuss data
associating SRPKs not only with additional mitotic
events, but also with other functional aspects of the
mammalian cell cycle.
Regulation of chromatin binding to the nuclear
envelope
Several macromolecular complexes are assembled by
various integral proteins of the nuclear envelope that
have been proposed to function as chromatin-anchor-
age platforms [74]. LBR is one of the key factors that
has been implicated in chromatin anchorage and was
shown to form oligomeric stuctures at the level of the
nuclear envelope [75–77]. The LBR–chromatin associa-
tion is probably mediated by electrostatic interactions
between the positively-charged residues of the N-termi-
nal domain of LBR and the negatively-charged phos-
phate groups of DNA [78]. The N-terminal domain of
LBR harbors a RS domain, the phosphorylation of
which not only reduces the positive charges, thereby
weakening the interaction with DNA, but also may
result in the disassembly of the oligomeric structure of
LBR [79]. The joint effect of the charge reduction and
the conformational change may render the phosphory-
lated monomeric N-terminal domains unable to anchor

the arrays of nucleosomes to the nuclear periphery. It
is well known that, during mitosis, the nuclear enve-
lope breaks down and chromosomes dissociate from
the inner nuclear membrane. Already at prophase,
binding of the membranous structures to chromosomes
is weakened. The RS domain of LBR is phosphoryla-
ted at the beginning of mitosis by nuclear-translocated
SRPK1 and potentially by Akt and Clk kinases that
may also target RS domains [26,36]. Furthermore, the
central mitotic kinase, cdk1, phosphorylates LBR at
Ser71 [80], which is located just upstream of the RS
repeats. It is therefore possible that these combinato-
rial phosphorylation events may result in chromosome
dissociation. This idea is consistent with a previous
study reporting that phosphorylation of LBR by mito-
tic extracts impairs chromatin association [81].
Fig. 2. A model illustrating the interactions between the NH
2
-terminal nucleoplasmic domain of LBR and P1 protamine. At the beginning of
spermiogenesis, the RS domain of LBR is unphosphorylated, allowing its association with phosphorylated protamine 1. LBR may act as a
docking site for the replacement of transition proteins (TP) by P1 protamine in certain chromatin layers that come close to the nuclear
periphery. Enzymes trapped in the inner nuclear membrane (INM) may also further modify the P1 protamine molecules, thereby facilitating
their deposition on sperm chromatin. The detachment of P1 protamine from the nuclear envelope and its tight binding to DNA is proposed
to occur through its dephosphorylation, whereas, at the same time, a similar dephosphorylation event may trigger the dissociation of TP
from sperm chromatin.
Serine-arginine protein kinases T. Giannakouros et al.
576 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
Regulation of chromatin reorganization during
G2 ⁄ M phase progression
Another mode of action of SRPK1 related to its

nuclear translocation at the G2 ⁄ M boundary involves
chromatin reorganization. A recent study demon-
strated that two SR proteins, SRp20 and ASF ⁄ SF2,
are released from mitotic chromosomes, during which
the H3 tail is modified at Ser10 by the activated aurora
kinase B, and reassociate with chromatin late in M
phase [82]. Hyperphosphorylation of these two SR
proteins by SRPK1 was also found to significantly
diminish their interaction with the H3 tail. Intrigu-
ingly, dissociation of ASF ⁄ SF2 from phosphorylated
histone H3 was required for the subsequent release of
HP1 (a key constituent of interphase hetechromatin)
from mitotic chromatin.
We propose that SRPKs (and potentially members
of the Akt and Clk family of kinases) may have a key
role at the beginning of mitosis by first mediating
the detachment of peripheral heterochromatin from
the inner nuclear membrane and, subsequently, the
removal of HP1, thus leading to chromosome conden-
sation (Fig. 3). An even more intriguing possibility is
that these phosphorylation events may also be applica-
ble during interphase for fine-tuning gene expression.
It has been suggested by Misteli [83] that the differen-
tial regulation of gene expression might involve the
inducible ‘potentiation’ of genomic loci, with subse-
quent displacement from their chromosome territory
and translocation to a transcriptionally silencing or
activating microenvironment. Because the coupling of
chromatin domains to the nuclear envelope has been
proposed to result in their transcriptional inactivation

[84], and HP1 proteins are well-known constituents of
‘silent’ chromatin, the regulated nuclear translocation
of SRPKs may contribute to the re-positioning and
‘unwinding’ of specific genomic loci, thus leading to
their transcriptional activation.
Regulation of cyclin transcription
SRPK2 has been implicated in the transcriptional
regulation of two members of the cyclin family. In
hematopoietic cells, SRPK2 was reported to enhance
cyclin A1 transcription [10], whereas, in neurons, it
was shown to trigger cell cycle progression and induce
apoptosis through regulation of cyclin D1 [47].
Cyclin A1 is a member of mammalian A-type cyclins
and is mainly expressed in male germ cells, being
essential for the passage of spermatocytes into meiosis
I [85]. In addition to male germ cells, elevated levels of
cyclin A1 expression have been detected in several leu-
kemic cell lines as well as in hematopoietic stem cells
and primitive precursors [86]. Up-regulation of cyclin
A1 by SRPK2 is accomplished through phosphoryla-
tion of the protein acinus that contains several RS
domains and its subsequent redistribution from nuclear
Fig. 3. Modulation of chromatin condensation at the beginning of mitosis by SR protein kinases. The combined phosphorylation of the RS
domain of LBR by nuclear translocated SRPK1 and the central mitotic kinase cdk1 (and potentially by Clk and Akt kinases) results in chromo-
some dissociation from the inner nuclear membrane. A concomitant combined phosphorylation event [i.e. phosphorylation of Ser10 of H3
by aurora B and phosphorylation of SR proteins ASF ⁄ SF2 and SRp20 (SR) by nuclear translocated SRPK1, and potentially by Clk and Akt
kinases] results in HP1 release from mitotic chromatin, further facilitating chromatin condensation.
T. Giannakouros et al. Serine-arginine protein kinases
FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS 577
speckles to the cytoplasm [10]. Acinus was originally

identified as a target of caspase-3, a cysteine protease
involved in activating chromatin condensation and
nuclear fragmentation during apoptosis; however, the
role of this protein during normal cellular growth has
not been determined [87,88]. Acinus is phosphorylated
by SRPK2 at Ser422, and acinus S422D, a SRPK2
phosphorylation mimetic, was shown to enhance cyclin
A1 transcription, whereas acinus S422A, an unphosph-
orylatable mutant, was shown to block the stimulatory
effect of SRPK2. Furthermore, siRNA-mediated
down-regulation of acinus or SRPK2 resulted in cyclin
A1 repression in leukemic cells and the cells were
arrested at the G1 phase. Interestingly, overexpressed
FLAG-SRPK1 was unable to associate with and phos-
phorylate recombinant acinus, indicating that the inter-
action between acinus and SRPK2 is specific [10]. To
our knowledge, this is the first report of two SRPK
family members exhibiting a differential recognition
pattern towards an RS domain-containing protein.
Cyclin D1 functions as a mitogenic sensor and is
one of the more frequently altered cell cycle regulators
in cancers [89]. It belongs to the family of mammalian
D-type cyclins that are G1-specific. Cyclin D1 associ-
ates with and allosterically activates CDK4 or CDK6,
thereby promoting restriction point progression during
the G1 phase [89]. Terminally differentiated neurons
are unable to reenter the cell cycle. Aberrant cell cycle
activation provokes neuronal cell death, whereas cell
cycle inhibition increases neuronal survival. SRPK2
triggers cell cycle progression in neurons and induces

apoptosis through regulation of nuclear cyclin D1 [47].
According to Jang et al. [47], up-regulation of cyclin
D1 in this system is not mediated through acinus phos-
phorylation but rather through inactivation of p53.
More specifically, it has been proposed that SRPK2
phosphorylates and activates SC35 and, thus, it may
inactivate p53 by blocking its phosphorylation at
Ser15 [47,90]. Interestingly, it has been also reported
that SC35 affects transcriptional elongation in a gene-
specific manner [91]. Thus, activation of SC35 may
lead to down-regulation of specific genes, including
p53. Because p53 represses cyclin D1 expression [92],
down-regulation of p53 may also result in cyclin D1
up-regulation.
In this respect, it should be noted that SRPKs have
been proposed to act as modifiers of the p53 pathway
in Drosophila (Patent WO ⁄ 2002 ⁄ 099427: SRPKs as
modifiers of the p53 pathway). More specifically, a
genetic screen identified that a SRPK mutation
enhanced cell death, as induced by the expression of
p53 in the Drosophila wing. Because Drosophila con-
tains more than one Srpk gene, it remains unclear
whether the regulation of p53 activity is exerted by a
specific SRPK (e.g. the SRPK2 homolog) and, more
importantly, whether this regulation is accomplished
solely through SC35.
SRPKs and metabolic signaling
The PGC-1 family of coactivators mediates various
environmental signals, thus regulating several meta-
bolic pathways in a tissue-specific manner [93]. Most

importantly, the PGC-1 coactivators play a critical role
in modulating glucose, lipid and energy homeostasis
that become deregulated in metabolic diseases such as
diabetes, obesity and cardiomyopathy. The first mem-
ber of the PGC-1 family, PGC-1a, was identified as a
cofactor for peroxisome proliferator activated recep-
tor c approximately one decade ago [94]. PGC-1a
activity is modulated by a large number of post-trans-
lational modifications, including phosphorylation by
several kinases, acetylation and deacetylation by
GCN5 and silent information regulator 1, respectively,
as well as O-GlcNAcylation by O-GlcNAc transferase
[95].
PGC-1a contains a RS domain that links insulin sig-
nal transduction to the repression of gluconeogenesis
[96]. This link is mediated through phosphorylation of
the RS domain that renders PGC-1a unable to coacti-
vate the forkhead transcription factor forkhead box
protein O1 (FOXO1), which is the main nuclear recep-
tor controlling the glyconeogenic program [96–98]. To
date, two kinases have been implicated in the phos-
phorylation of the RS domain: Akt2 and Clk2 [97,98].
Akt2 phosphorylates only Ser570, which is the last ser-
ine in the first repeat of RS dipeptides (RSRSR
SFSR)
[97], whereas Clk2 probably phosphorylates the entire
RS domain [98].
A central question that arises is whether PGC-1a is
also phosphorylated by members of the SRPK family,
and whether this phosphorylation can repress its tran-

scriptional activity. It was previously shown that
SRPK1 can phosphorylate in vitro the RS domain of
PGC-1a [79], although a similar phosphorylation event
has not yet been shown to occur in vivo. We anticipate
that this type of phosphorylation may also take place
in vivo and not only by SRPK1, but also by other
members of the SRPK family, to an extent propor-
tional to the expression levels of SRPKs in liver.
Another important issue is the response to insulin.
Akt2 is an insulin-responsive kinase, whereas it was
shown to phosphorylate Clk2 at Thr343, leading to an
increase of Clk2 protein stability and therefore activity
[98]. Clk2 was therefore suggested to function as
an insulin-induced gluconeogenic repressor. Yet Akt
Serine-arginine protein kinases T. Giannakouros et al.
578 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
kinases can also phosphorylate SRPK2 at Thr492 and
mediate its nuclear translocation [47], thus making
SRPK2 an insulin-responsive kinase as well. It is still
unknown whether insulin has any effect on SRPK1
and ⁄ or on SRPK1a, either directly through phosphor-
ylation by Akt kinases or through another indirect sig-
nalling mechanism. In this respect, it should be noted
that SRPK1a contains two LXXLL motifs [16] that
are assumed to facilitate the interaction of different
proteins with nuclear receptors. All these phosphoryla-
tion events may act in a complementary fashion
(Fig. 4A), thus constituting a fine-tuning mechanism
that modulates the interaction of PGC-1a with various
transcription factors and allows the expression of

specific gene sets in different physiological settings.
PGC-1a also stimulates mitochondrial biogenesis
through coactivation of nuclear respiratory factor-1
(NRF-1) [99]. Indeed, PGC-1a expression in both mus-
cle and fat cells activates the expression of several
genes of the oxidative phosphorylation pathway,
including cytochrome c oxidase subunits II and IV,
and ATP synthase. Although the RS domain of PGC-
1a mediates its interaction with FOXO1, the 200-400
amino acid region of PGC-1a is responsible for the
Fig. 4. Regulation of PGC-1a transcriptional activity by its RS region (A) Akt2, Clk2 and potentially SRPK2 phosphorylate various serine resi-
dues within the RS region of PGC-1a, thus impairing its interaction with FOXO-1 to a different extent each time. Akt2 phosphorylates only
Ser570 (purple), SRPK2 (and possibly other SRPKs as well) may phosphorylate the serines within the three repeats of RS dipeptides (blue),
whereas Clk2 probably phosphorylates the entire RS domain (red). Akt2 is activated by insulin, whereas it phosphorylates Clk2 at Thr343,
leading to an increase of Clk2 protein stability and activity. Akt kinases also phosphorylate SRPK2 at Thr492 and mediate its nuclear translo-
cation, thus rendering SRPK2 molecules available to phosphorylate nuclear PGC-1a. It is not clear yet whether there is further cross-regula-
tion between SRPK, Clk and Akt kinases. The stoichiometry of PGC-1a phosphorylation (i.e. the number of protein molecules per cell that
are phosphorylated) and also the exact serines of the RS region that are modified in each molecule may constitute a fine-tuning mechanism
that regulates the transcription of gluconeogenic genes and mediates PGC-1a responsiveness to insulin. (B) A more permanent inactivation
of the RS domain (in muscle and fat cells) may be achieved through binding of p32 protein. A central function of p32 protein is to associate
with and impair the phosphorylation of RS domains. p32 protein may obstruct the interaction of PGC-1a with FOXO-1 that requires the RS
domain, thus allowing the available PGC-1a molecules to interact with NRF-1 and promote the transcription of specific genes involved in
oxidative phosphorylation.
T. Giannakouros et al. Serine-arginine protein kinases
FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS 579
PGC-1a ⁄ NRF-1 interaction. Thus, the ability of PGC-
1a to interact with different transcription factors
allows for the coordinated expression of gene sets in
specific cellular contexts and in response to specific sig-
nals. A recent study adds another piece to the puzzle

and further strengthens the hypothesis that SRPKs
may be actively involved in the phosphorylation of
PGC-1a. Fogal et al. [100], extending previous obser-
vations [101], reported that p32 protein plays a decisive
role in maintaining mitochondrial oxidative phosphor-
ylation. Knocking down p32 expression in human can-
cer cells resulted in a reduced expression of oxidative
phosphorylation-related polypeptides and shifted the
cell metabolism from oxidative phosphorylation to gly-
colysis. p32 is an ‘all-around’ cellular protein found in
the nucleus, cytoplasm, mitochondria and cell surface
with essentially unknown physiological role(s) [102].
Yet p32 was reported to bind the RS domains of both
ASF ⁄ SF2 and LBR and inhibit the phosphorylation of
these molecules by SRPKs [67,103,104]. Even though it
remains to be proven, we speculate that p32 protein
drives PGC-1a activity toward specific gene sets
involved in oxidative phosphorylation by obstructing
its interaction with FOXO1, thus allowing the avai-
lable molecules to interact with NRF-1 (Fig. 4B).
Regulation of the SRPK family
members
SRPKs have been considered to be constitutively
active kinases because the expression of SRPK family
members in bacteria, which lack the post-translational
modification machinery of eukaryotic cells, has shown
that they are able to efficiently phosphorylate their
substrates [26,29,105]. Furthermore, co-expression of
SRPK1 and its substrate SF2 ⁄ ASF in Escherichia coli
results in the phosphorylation and splicing activity of

the latter [106]. In support of these findings, structural
and biochemical studies on SRPK1 and Sky1p have
shown that their kinase core domains can adopt,
through a network of connections, an active conforma-
tion even when they are extensively mutated or their
N-terminal and spacer domains are truncated, provid-
ing more evidence of their constant activity and sub-
strate phosphorylation mechanisms [107,108].
It might be expected that this family of kinases is
somehow regulated. Reports from various studies
agree that the determining factor in SRPK regulation
is their subcellular partitioning. It has been demon-
strated that SRPKs are primarily located in the cyto-
plasm of interphase cells and, to a lesser extent, in the
nucleus [4,9,16,37]. This cytoplasmic sequestration
mainly depends on the existence of the unique and
divergent spacer sequence in each family member that
divides the conserved catalytic kinase domains into
two halves. As initially reported for Dsk1 and Sky1p
in S. pombe and S. cerevisiae, respectively, deletion of
the spacer domain results in their nuclear transloca-
tion, with no apparent loss of activity. Studies have
shown that this sequence acts like a cytoplasmic
anchor critical for SRPK regulation because the con-
stant accumulation of mutant Sky1p in the nucleus
provokes inhibition of cell growth [3,11]. In addition,
removal of the spacer domain in SRPK1 ⁄ 2 forces their
displacement to the nucleus, causing the aggregation of
splicing factors and possibly affecting gene expression
[4]. The cytoplasmic anchoring of SRPKs has recently

been shown to be mediated by their association with
specific members of molecular chaperones (Fig. 1).
Thus, direct interaction of SRPK1 with cochaperones
Aha1 and heat shock protein Hsp40 mediates the for-
mation of a complex with the Hsp70 ⁄ Hsp90 machinery
[46]. Furthermore, SRPK2 directly associates with the
-b and -e isoforms of 14-3-3 family of proteins in an
Akt phosphorylation-depended manner in the cyto-
plasm of neuronal cells [47].
The interaction of SRPK1 with the molecular chap-
erones could be modulated by signal(s) resulting in the
release and subsequent translocation of the kinase to
the nucleus. One option is that SRPKs may be post-
translationally modified in response to signaling. In
this respect, a previous study indicated that SRPK1 is
phosphorylated and partially activated by casein
kinase 2 (CK2) [109]. However, it remains to be deter-
mined whether CK2 has any effect on the nuclear
translocation of the kinase. Furthermore, Akt was pro-
posed to mediate the nuclear translocation of SRPK2
by phosphorylating it at Thr492, whereas 14-3-3 mole-
cules were shown to interact with Akt-phosphorylated
SRPK2 and inhibit its nuclear translocation [47]. Of
note, the major CK2 phosphorylation site (
SDDD,
Ser51 in human SRPK1) is conserved among SRPK
family members, whereas the Akt site (HDRSR
TVS,
Thr492 in human SRPK2) is not, and probably repre-
sents a SRPK2-specific mode of regulation. A second

option is that the nuclear translocation of the kinases
may be accomplished through the reorganization
and ⁄ or modification of specific components of the
chaperone complex. Supporting this option is the dem-
onstration that inhibition of the ATPase activity of
Hsp90 with 17-AAG results in partial translocation of
SRPK1 in the nucleus [46]. Finally, as already noted,
the ICP27 protein of HSV-1 interacts with SRPK1 and
promotes its translocation to the nucleus [51]. Yet
none of the above signals is cell cycle-regulated; there-
fore, the signal triggering the nuclear translocation of
Serine-arginine protein kinases T. Giannakouros et al.
580 FEBS Journal 278 (2011) 570–586 ª 2011 The Authors Journal compilation ª 2011 FEBS
SRPK1 at the G2 ⁄ M boundary remains to be
determined.
The molecular mechanism that mediates the nuclear
export of SRPKs also remains obscure. Only the fis-
sion yeast Dsk1 has been shown to harbor an active
nuclear export signal inside its spacer region that is
recognized by CRM1 because Dsk1 export is impaired
by leptomycin B treatment [110]. Functional nuclear
export signal sequences that may explain efficient
transport through the nuclear membrane remain to be
identified in other SRPK family members.
In addition to subcellular partitioning, other modes
of SRPK regulation also exist. Despite the fact that
unmodified kinase molecules are active, it was shown
that CK2-mediated phosphorylation of SRPK1
enhances its activity by six-fold [109]. The primary
CK2 target site (Ser51 at SRPK1) was also found to

be modified in vivo by MS in kinome-wide phospho-
proteomics studies in HeLa cells [111–113]. These
reports also indicate the existence of additional
phosphorylation sites on SRPK molecules (http://
www.phosphosite.org/homeAction.do; keywords
‘SRPK1’ and ‘SRPK2’), suggesting that there is more
than one unidentified signal, which could affect either
their activity or shuttling through the modulation of
their interaction with other proteins.
Another aspect of the modulation SRPK activity
involves transient interaction with nonshuttling protein
complexes. It was recently shown that both SRPK1
and SRPK1a could directly interact with SAFB1 and
SAFB2, albeit with different affinities. This association
does not depend on the spacer domain, as was shown
for other protein complexes of SRPK1, but rather on
the N-terminal and core kinase domains. Interaction
with SAFB molecules impaired the catalytic activity of
SRPK1 ⁄ 1a, whereas the nuclear subfraction of the
kinases, which was found to be associated with the
nuclear matrix via SAFB proteins, was inactive [17].
Given that SAFB proteins are also sequestrated in
stress-induced subnuclear bodies, along with splicing
factors and RNA molecules in response to stress
[114,115], it is intriguing to consider that their interac-
tion with SRPK1 ⁄ 1a and subsequent inactivation of
the kinases could provide an additional mechanism of
controlling SRPK activity when the cell needs to react
promptly to a variety of signals.
Conclusions

In conclusion, the accumulated knowledge on SRPK
function not only enlightens many aspects of their
influence on fundamental cellular mechanisms, but also
raises questions that need to be addressed in future
studies to obtain insight into their role in the cell.
These include:
l
The conformational changes induced by the phos-
phorylation of the RS domain.
l
The exact share the SRPKs hold amongst the other
kinases that also recognize RS dipeptides, such as the
Clk and Akt family of kinases, as well as the cross-reg-
ulation between them.
l
The specific role(s) of each one of the SRPK family
members and the significance of the ‘spacer domain’
for the functional properties and the particular regula-
tion of each kinase.
l
The extra- and intracellular signals that regulate
SRPK function.
Further investigations into the above issues, in light
of a thorough examination of the evolutionary history
of the SRPK genes, will help to unveil the functional
presence of the SRPK family in cellular life.
Acknowledgements
We thank John Georgatsos for critically reading the
manuscript. This work was supported by grants from
the Greek General Secretariat of Research and Tech-

nology and the Greek Ministry of Education.
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