Cap-binding complex (CBC)

Abstract

The 7mG (7-methylguanosine cap) formed on mRNA is fundamental to eukaryotic gene expression. Protein complexes recruited to 7mG mediate key processing events throughout the lifetime of the transcript. One of the most important mediators of 7mG functions is CBC (cap-binding complex). CBC has a key role in several gene expression mechanisms, including transcription, splicing, transcript export and translation. Gene expression can be regulated by signalling pathways which influence CBC function. The aim of the present review is to discuss the mechanisms by which CBC mediates and co-ordinates multiple gene expression events.

Figure 1. 7mG structure

The 7mG structure is depicted including the first and second transcribed nucleotides. Cap 2 structure is depicted, i.e. methylated on the O-2 position of ribose on the first and second transcribed nucleotides.

Figure 2. 7mG synthesis in S. cerevisiae, Schizosaccharomyces pombe and humans

The RNA triphosphatase enzymes are depicted in light blue, the RNA guanylyltransferase in dark blue and the RNA guanine-7-methyltransferases in green. (a) In S. cerevisiae, the RNA guanylyltransferase (Ceg1p) and the RNA triphosphatase (Cet1p) interact with the Ser⁵-phosphorylated RNA pol II CTD via the Ceg1p subunit [–172]. The RNA guanine-7-methyltransferase (Abd1p) is recruited to the transcription initiation site via an interaction with phosphorylated RNA pol II CTD [12,95,173]. (b) In S. pombe, the RNA triphosphatase (Pct1p) and the RNA guanylyltransferase (Pce1p) independently interact with phosphorylated RNA pol II CTD [171,174]. Pct1p interacts with DSIF (DRB sensitivity-inducing factor) (Spt4/Spt5) and P-TEFb (Cdk9/Cyclin T1) [175,176]. Pce1p interacts with DSIF [176,177]. The RNA guanine-7-methyltransferase (Pcm1p) functions at the transcription initiation site, but does not physically associate with RNA pol II. Pcm1p and P-TEFb are in a complex and Pcm1p is required for P-TEFb recruitment to chromatin and transcription elongation [11,178,179]. (c) In humans, the RNA triphosphatase and the RNA guanylyltransferase activities reside in the same polypeptide, RNGTT. RNGTT interacts with DSIF (Spt4/5), which stimulates its activity up to 5-fold [13,180]. RNGTT promotes transcription elongation independently of its catalytic activity by overcoming NELF-dependent transcriptional pausing [13]. RNGTT is recruited to transcription initiation sites via an interaction with Ser⁵-phosphorylated RNA pol II CTD, which stimulates guanylyltransferase activity [169,181,182]. The RNA guanine-7-methyltransferase complex (RNMT/RAM) interacts with RNGTT and indirectly with RNA pol II [5,183]. RNMT also interacts with importin-α (Imp-α), which stimulates cap methyltransferase activity [184].

Figure 3. CBC functions

CBC, composed of Cbp20 and Cbp80 subunits, binds to 7mG located at the 5′ end of RNA pol II transcripts. CBC interacts with a spectrum of factors mediating RNA metabolism and translation mechanisms. CycT1, cyclin T1; Dcp, decapping mRNA; DXO, decapping exoribonuclease; Xm2, 5′–3′ exoribonuclease 2.

Figure 4. CBC functions in eukaryotic gene expression

(a) CBC is required for pre-mRNA processing. The co-transcriptional binding of CBC to 7mG prevents the decapping activities of pre-mRNA degradation complexes [DXO (decapping exoribonuclease) and Dcp (decapping mRNA) Xrn2 (5′–3′ exoribonuclease 2)] and promotes pre-mRNA processing. CBC recruits P-TEFb [Cdk9/Cyclin T1 (CycT1)] to transcription initiation sites of specific genes promoting phosphorylation of the RNA pol II CTD at Ser² residues. This results in the recruitment of splicing factors including SRSF1, which regulates both constitutive and alternative splicing events. Furthermore, CBC interacts with splicing machinery components that results in the spliceosomal assembly. CBC interacts with NELF and promotes pre-mRNA processing of replication-dependent histone transcripts. (b) CBC forms a complex with Ars2 and promotes miRNA biogenesis by mediating pri-miRNA processing. (c) CBC/Ars2 promotes pre-mRNA processing of replication-dependent histone transcripts. (d) CBC promotes export of U snRNA. CBC interacts with PHAX, which recruits export factors including CRM1 and RAN·GTP. (e) CBC promotes export of mRNA. For export of transcripts over 300 nucleotides, hnRNP C interacts with CBC and inhibits the interaction between CBC and PHAX, allowing the CBC to interact with TREX and the transcript to be translocated to the cytoplasm. CBC interacts with the PARN deadenylase and inhibits its activity, protecting mRNAs from degradation. (f) CBC mediates the pioneer round of translation. Cbp80 interacts with CTIF, which recruits the 40S ribosomal subunit via eIF3 to the 5′ end of the mRNA for translation initiation. Upon binding of importin-β (Imp-β) to importin-α (Imp-α), mRNA is released from CBC and binds to eIF4E for the initiation of the standard mode of translation. CBC-bound mRNP components not found in eIF4E-bound mRNPs are CTIF, exon junction complex (EJC) and PABPN1. (g) The standard mode of translation is mediated by eIF4E cap-binding protein. eIF4E is a component of the eIF4F complex which promotes translation initiation.