RNA UK 2008
Regulation of gene expression by the RNA-binding
protein Sam68 in cancer
Prabhakar Rajan*†1 , Luke Gaughan†, Caroline Dalgliesh*, Amira El-Sherif‡, Craig N. Robson†, Hing Y. Leung§ and
David J. Elliott*
*Institute of Human Genetics, Newcastle University, Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, U.K., †Northern Institute for Cancer
Research, Newcastle University, Newcastle upon Tyne NE2 4HH, U.K., ‡Department of Pathology, Royal Victoria Infirmary, Newcastle upon Tyne NE4 6BE,
U.K. and §Beatson Institute for Cancer Research, Glasgow G61 1BD, Scotland, U.K.
Abstract
Sam68 (Src-associated in mitosis 68 kDa) is the prototypical member of the STAR (signal transducer and
activator of RNA) family of RNA-binding proteins. Sam68 is implicated in a number of cellular processes
including signal transduction, transcription, RNA metabolism, cell cycle regulation and apoptosis. In the
present review, we summarize the functions of Sam68 as a transcriptional and post-transcriptional regulator
of gene expression, with particular relevance to cancer.
Introduction
Sam68 (Src-associated in mitosis 68 kDa) belongs to the
STAR (signal transducer and activator of RNA) family
of RNA-binding proteins that link signalling pathways to
RNA processing (for a review, see [1]). STAR family proteins
are characterized by at least three functional motifs: a single
extended KH [hnRNP (heterogeneous nuclear ribonucleoprotein) K homology] domain embedded within a larger
conserved GSG (GRP33/Sam68/GLD1) domain consisting
of approx. 80- and 30-amino-acid sequences referred to as
the NK (N-terminus of KH) and CK (C-terminus of KH)
regions respectively (Figure 1).
The RNA-binding ability of Sam68 is harboured within
the GSG domain, which is required for homodimerization
and sequence-specific binding to RNA targets. The GSG
domain is flanked by a proline-rich WW domain (protein–
protein interaction domain containing two conserved tryptophan residues)- and SH (Src homology domain) 3-binding
regions, and SH2-interacting tyrosine-rich motifs, which
mediate interplay with numerous cell signalling components
in response to different stimuli and critically regulate Sam68
function (reviewed in [1]).
Sam68 and RNA processing
A major role for Sam68 is in the regulation of RNA metabolism through RNA processing and export. Sam68 binds nonspecifically to poly(U) and poly(A) RNA, and specifically
to the high-affinity binding sequences UAAA or UUUA
Key words: cancer, cell cycle, RNA processing, signal transducer and activator of RNA (STAR),
Src-associated in mitosis 68 kDa (Sam68), transcription.
Abbreviations used: AR, androgen receptor; ER, oestrogen receptor; ERK, extracellular-signalregulated kinase; hnRNP, heterogeneous nuclear ribonucleoprotein; KH, hnRNP K homology;
MMTV, murine-mammary-tumour virus; CK, C-terminus of KH; NK, N-terminus of KH; RNAPII,
RNA polymerase II; Sam68, Src-associated in mitosis 68 kDa; GSG, GRP33/Sam68/GLD1; SH,
Src homology domain; STAR, signal transducer and activator of RNA; SWI/SNF, mating-type
switch/sucrose non-fermenting; SUMO, small ubiquitin-related modifier.
1
To whom correspondence should be addressed (email [email protected]).
Biochem. Soc. Trans. (2008) 36, 505–507; doi:10.1042/BST0360505
as determined by SELEX (systematic evolution of ligands
by exponential enrichment) [2]. The 3 -UTR (3 -untranslated
region) contains an AU-rich sequence, and recent work by
our group suggests that this region is a target for Sam68
binding (Y. Liu and D.J. Elliott, unpublished work). Although
a direct role in translation has not been demonstrated, Sam68
has been implicated in modulating the nuclear export and
cytoplasmic utilization of viral mRNAs [3,4].
Despite a predominantly nuclear localization, Sam68 was
first identified as the major phosphorylation substrate for Src
in the cytoplasm during mitosis. Post-translational modifications can affect Sam68 function, by critically regulating
intracellular localization and accessibility to RNA: tyrosine
residues are phosphorylated by non-receptor tyrosine kinases (such as Src and Fyn) and subsequently bound by SH2
domains preventing association with RNA [5]; methylation
of RG (arginine/glycine)-rich regions by arginine N-methyltransferases is required for RNA export [4]; acetylation of
lysine residues within the GSG domain by histone acetyltransferases positively regulates RNA binding [6]; and
SUMO (small ubiquitin-related modifier) modification
enhances the ability of Sam68 to repress cyclin D1 expression
[7].
The above findings suggest that Sam68 behaves as a
multifunctional adaptor by linking signalling pathways to
the transcriptional and post-transcriptional regulation of gene
expression. In particular, Sam68 was shown to induce ERK
(extracellular-signal-regulated kinase)-mediated inclusion
of CD44 variable exon v5 in response to Ras activation
by phorbol ester stimulation [8]. Sam68 was identified as a
target for phosphorylation by ERK and was found to bind
exonic splice-regulatory elements within CD44 exon v5.
Misregulation of CD44 splice variants is implicated in cancer.
More recently, Sam68 was shown to associate with Brm,
a component of the SWI/SNF (mating-type switch/sucrose
non-fermenting) chromatin remodelling complex, which
also favours inclusion of CD44 variant exons [9]. The study
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Figure 1 Sam68 domain structure
Figure 2 A model for Sam68 function in the regulation of gene
Tripartite GSG domain containing a KH domain, NK region and CK region.
Flanking domains contain six consensus proline-rich motifs (P0–P5),
expression
Distinct functions for Sam68 protein (i) at the promoter with a role
RG-rich sequences, and a tyrosine (Y)-rich region close to the C-terminal
NLS (nuclear localization sequence).
in transcriptional co-regulation; and (ii) associated with the C-terminal
domain (CTD) of RNAPII, with a role in RNA processing. These functions
may be regulated by the post-translational modifications such as
phosphorylation (P). MAPK, mitogen-activated protein kinase; RTK,
receptor tyrosine kinase; TF, transcription factor.
suggested that Brm and Sam68 may co-operate to slow the
rate of RNAPII (RNA polymerase II) on genes regulated
by SWI/SNF, thus facilitating recruitment of the splicing
machinery to suboptimal splice sites within variable exons
[9]. In this way, Sam68 may contribute to cross-talk between
cell signalling, transcription and RNA processing.
Sam68 and transcriptional regulation of
gene expression
Direct evidence implicating Sam68 in transcriptional regulation of gene expression is limited: binding of Sam68 to hnRNP
K inhibits the function of hnRNP K in transcriptional
activation of a reporter driven by the CT promoter element
of the proto-oncogene c-myc, through direct protein–protein
interaction [10]. Sam68 has been noted to repress various
mammalian and viral promoter constructs, and potently
inhibit the transcriptional activity of the multifunctional
adaptor CBP [CREB (cAMP-response-element-binding protein)-binding protein] in a GAL4-dependent reporter system
independent of its RNA-binding ability [11]. In these
scenarios, it is thought that Sam68 may behave as a competitive inhibitor of positive regulators of transcription.
The role of Sam68 as a transcriptional co-regulator is
likely to be promoter- and transcription-factor-dependent:
Sam68 has been shown to enhance the transcriptional activity
of the AR (androgen receptor), a nuclear hormone receptor
transcription factor [12]. This observed effect was seen using
a reporter system driven by the steroid hormone-responsive
MMTV (murine-mammary-tumour virus) promoter, and
was independent of the ability of Sam68 to bind RNA.
AR transcription function is important in prostate cancer
progression, and Sam68 has been found to be overexpressed
in prostate tumours [12,13].
The co-activator function of Sam68 may also extend to
other nuclear hormone receptors: Sam68 haploinsufficiency
has been shown to delay the onset of mammary tumorigenesis
driven by the polyoma middle T-antigen oncogene under
transcriptional control of the MMTV promoter [14]. Since
ER (oestrogen receptor) signalling is important in breast
cancer and the MMTV promoter is also ER-dependent,
Sam68 may also be acting as a co-activator of ER-dependent
transcription in mammary development and tumorigenesis.
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The evidence implicating Sam68 in cancer is contradictory:
Sam68 was initially thought to be a tumour suppressor, but
more recently it has been shown to exhibit a pro-oncogenic
function (reviewed in [15]). On the one hand, Sam68 overexpression induces G1 growth arrest, and down-regulation
of cyclins D1 and E independently of RNA-binding capacity
[16]. Sam68 SUMOylation on lysine residues also enhances
this ability to repress cyclin D1 expression [7]. However, on
the other hand, Sam68 knockdown in prostate cancer cells
delays cell cycle progression and reduces proliferation, but
causes an accumulation of cyclin D1 [13].
Sam68 is a downstream target of important signalling pathways activated in carcinogenesis such as receptor and nonreceptor tyrosine kinases and RET (rearranged during transfection) pathways (reviewed in [15]). Since ER signalling and
AR signalling are important in breast and prostate cancers
respectively, Sam68 may be functioning as a steroid hormone
receptor transcriptional co-activator in these tumours. Sam68
may also have a role in progression of these cancers to a
hormone-refractory phenotype.
Conclusions
On the basis of current evidence, it is unclear whether Sam68
is a tumour suppressor or classical oncogene. Sam68 appears
to have biochemically distinct roles in the cell depending on
post-translational modifications as a result of the activation
of different signalling pathways in cancer (Figure 2). Further
RNA UK 2008
studies are required to clearly define the functional roles
of Sam68 in cancer with the assistance of tissue-specific
genetically engineered models to study autochthonous
tumours.
We acknowledge the following sources of funding: the Medical
Research Council, Research Councils UK and the Association for
International Cancer Research.
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Received 21 January 2008
doi:10.1042/BST0360505
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