The human synMuv-like protein LIN-9 is required for transcription of G2/M genes and for entry into mitosis

Abstract

Regulated gene expression is critical for the proper timing of cell cycle transitions. Here we report that human LIN-9 has an important function in transcriptional regulation of G2/M genes. Depletion of LIN-9 by RNAi in human fibroblasts strongly impairs proliferation and delays progression from G2 to M. We identify a cluster of G2/M genes as direct targets of LIN-9. Activation of these genes is linked to an association between LIN-9 and B-MYB. Chromatin immunoprecipitation assays revealed binding of both LIN-9 and B-MYB to the promoters of G2/M regulated genes. Depletion of B-MYB recapitulated the biological outcome of LIN-9 knockdown, including impaired proliferation and reduced expression of G2/M genes. These data suggest a critical role for human LIN-9, together with B-MYB, in the activation of genes that are essential for progression into mitosis.

A Lin-9 complex is recruited by B-Myb to activate transcription of G2/M genes in undifferentiated embryonal carcinoma cells

“…Significantly, the expression of genes regulated at the other cell-cycle stages was not diminished; Cyclin E2 that is required for G 1 /S transition was upregulated whereas B-Myb whose expression peaks during S-phase was unaffected (Figure 2b). Interestingly, Cyclin A2 and Cdc2 expression was not significantly changed by depletion of Lin-9 in F9 cells, although previous studies in adult cells showed transcriptional dependence on Lin-9 (Osterloh et al, 2006). B-Myb has been found to be required to remove a repressive E2F4 complex at the Cdc2 promoter (Zhu et al, 2004), and possibly the lack of this repressor in F9 cells makes this activity redundant.…”

“…Specific enrichment was calculated relative to input chromatin levels using % of input ¼ 2 DCt , where DCt ¼ Ct 1% input -Ct IP . Re-ChIP experiments were performed as described by Osterloh et al (2006). Each ChIP experiment was carried out on biological triplicates.…”

“…In S-G 2 , LINC switches to B-Myb to activate genes required for G 2 /M transition and mitosis (Schmit et al, 2007). Depleting Lin-9 from human cells by RNA interference delayed the G 2 /M transition (Osterloh et al, 2006;Pilkinton et al, 2007b) and downregulated several genes required for entry into mitosis, spindle assembly and exit from mitosis (Osterloh et al, 2006). Reporter assays have shown that Lin-9 and B-Myb coactivate transcription of G 2 /M genes and chromatin immunoprecipitations (ChIP) have located Lin-9 and B-Myb on their promoters (Osterloh et al, 2006;Schmit et al, 2007;Pilkinton et al, 2007b).…”

Loss of LIN9, a member of the DREAM complex, cooperates with SV40 large T antigen to induce genomic instability and anchorage-independent growth

“…The DREAM (or LINC) complex is a master regulator of mitotic gene expression during the cell cycle (Schmit et al, 2007, reviewed in Blais and Dynlacht, 2007;Litovchick et al, 2007;Osterloh et al, 2007;Pilkinton et al, 2007a;Knight et al, 2009). Several proteins that are involved in chromosome segregation and cytokinesis, such as Bub1 and survivin, are direct targets of DREAM (Osterloh et al, 2007;Schmit et al, 2007;Pilkinton et al, 2007b;Knight et al, 2009).…”

“…Several proteins that are involved in chromosome segregation and cytokinesis, such as Bub1 and survivin, are direct targets of DREAM (Osterloh et al, 2007;Schmit et al, 2007;Pilkinton et al, 2007b;Knight et al, 2009). DREAM consists of a fiveprotein core module and associated proteins (Litovchick et al, 2007;Schmit et al, 2007;Pilkinton et al, 2007a;Knight et al, 2009).…”