Cyclin L2, a Novel RNA Polymerase II-associated Cyclin, Is Involved in Pre-mRNA Splicing and Induces Apoptosis of Human Hepatocellular Carcinoma Cells

Abstract

We report the cloning and functional characterization of human cyclin L2, a novel member of the cyclin family. Human cyclin L2 shares significant homology to cyclin L1, K, T1, T2, and C, which are involved in transcriptional regulation via phosphorylation of the C-terminal domain of RNA polymerase II. The cyclin L2 protein contains an N-terminal ''cyclin box'' and C-terminal dipeptide repeats of alternating arginines and serines, a hallmark of the SR family of splicing factors. A new isoform and the mouse homologue of human cyclin L2 have also been cloned in this study. Human cyclin L2 is expressed ubiquitously in normal human tissues and tumor cells. We show here that cyclin L2 co-localizes with splicing factors SC-35 and 9G8 within nuclear speckles and that it associates with hyperphosphorylated, but not hypophosphorylated, RNA polymerase II and CDK p110 PITSLRE kinase via its N-terminal cyclin domains. It can also associate with the SC-35 and 9G8 through its RS repeat region. Recombinant cyclin L2 protein can stimulate in vitro mRNA splicing. Overexpression of human cyclin L2 suppresses the growth of human hepatocellular carcinoma SMMC 7721 cells both in vitro and in vivo, inducing cellular apoptosis. This process involves up-regulation of p53 and Bax and decreased expression of Bcl-2. The data suggest that cyclin L2 represents a new member of the cyclin family, which might regulate the transcription and RNA processing of certain apoptosis-related factors, resulting in tumor cell growth inhibition and apoptosis.Cyclins are key regulatory proteins that complex with and activate cyclin-dependent kinase (CDK) 1 subunits (1-4), playing pivotal roles in the regulation of cell cycle progression (5).Cyclins may be currently classified into two major groups, seemingly reflecting their functions. The cell cycle regulator cyclins, composed primarily of cyclin A, B, D1, D2, D3, E, and F, function with their CDK partners including CDK1, -2, -3, and -4 to regulate promotion of the cell cycle. The transcription regulator cyclins, which include cyclin C, H, K, L1, T1, and T2, work together with CDK7, -8, and -9 and tend to play roles in transcriptional regulation (6). The proposed functions of cyclin G 1 and G 2 are seemingly distinct from either cell cycle or transcriptional regulation (7). Other cyclins have also been identified, but their roles in cell cycle control and RNA transcription, if any, are unclear and remain to be studied.Transcription regulator cyclin/CDK pairs are associated with the initiation, elongation, and processing of RNA transcripts. During this process cyclins interact with the large subunit of RNA polymerase II, phosphorylating the C-terminal domain (CTD) via their partner CDKs. It is widely accepted that transcription and RNA processing are closely related. Capping enzymes, polyadenylation factors, and splicing factors assemble at the CTD of RNA polymerase II. Hyperphosphorylated RNA polymerase II (RNAPIIo) stimulates RNA splicing, and the CTD has been observed to target some splicing factors to R...

Amplification of Cyclin L1 is associated with lymph node metastases in head and neck squamous cell carcinoma (HNSCC)

“…Up to now, only little is known about its function. CCNL1, like the closely related protein CCNL2, contain a N-terminal cyclin box but differs from the other members of the cyclin family by the presence of a so-called RSdomain, which is a hallmark of proteins involved in pre-mRNA processing (de Graaf et al, 2004;Yang et al, 2004). Both proteins were recently found to be colocalised with the splicing factor SC35 within nuclear speckles.…”

Identification of novel cyclin gene fusion transcripts in endometrioid ovarian carcinomas

“…Yang et al 20. showed a similar tumor suppressor activity of Cyclin L2 in hepatocarcinoma and that both cyclin domains were fundamental for the protein's proper functioning 20. The fusion PCMTD1‐CCNL2 leads to a chimeric Cyclin L2α lacking the first cyclin domain (76–150) and containing only 27 aa of the second cyclin domain (Figure 1).…”

“…The CCNL2 gene encodes three cyclin L2 isoforms 19. The main isoform (Cyclin L2α) contains two cyclin domains, spanning amino acids 76–150 and 192–281, and a C‐terminal RS site (arginine–serine dipeptide) (385–423) that plays a role in protein–protein interactions with the SR family of splicing factors 20. The three splicing variants L2βA1/2/3 have exon 6 as the last coding exon and code for the 226 aa Cyclin L2βA isoform.…”

“…Cyclin L2, which is different from most other cyclins, is expressed during the entire cell cycle and was detected in many tissues, ovary included (Table 2). Cyclin L2 participates together with Cyclin L1 and CDK11 in pre‐mRNA splicing processes 19, 20. Lack of a functional Cyclin L2 may impair normal splicing mechanisms as all three Cyclin L2 isoforms have been shown to be fundamental components of the splicing complex 19…”

“…showed that it acts as a tumor suppressor protein in gastric cancer enhancing both apoptosis and chemosensitivity. Yang et al 20. showed a similar tumor suppressor activity of Cyclin L2 in hepatocarcinoma and that both cyclin domains were fundamental for the protein's proper functioning 20.…”