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The classical work by Price laboratory on the identification of P-TEFb in late 1990’s, and Zhou and Bensaude laboratories on the inhibition of P-TEFb orchestrated by the non-coding 7SK snRNA in the fall of 2001 have laid the groundwork for our understanding of regulating Pol II pause release in multicellular organisms.
Since then, intensive research by many investigators has ensued, revealing the widespread control of gene transcription at the elongation step and critical importance for P-TEFb in promoting Pol II elongation. At the same time, the composition of 7SK snRNP as well as many fundamental principles governing the sequestration and inhibition of P-TEFb within the inhibitory complex have been worked out. Our recently reported review article provides a historical perspective on how the field has developed. Moreover, we have summarized recent progress describing fascinating mechanisms that tether 7SK snRNP to chromatin as well as those that release P-TEFb from 7SK snRNP (Table 1).
Table 1. Summary of mechanisms that direct the anchoring and disintegration of canonical 7SK snRNP on chromatin.
What emerges from recent developments is that 7SK snRNP is not an inert and isolated particle roaming the nucleoplasm and harboring inactive P-TEFb. Rather, it can be contacted by numerous auxiliary factors: while chromatin adaptor factors (Ch-AFs) deliver 7SK snRNP to gene promoters or enhancers, P-TEFb-release factors (P-REFs) stimulate P-TEFb activation at appropriate circumstances, for a controlled gene-specific transition of Pol II from pausing into productive elongation. In turn, we propose a unifying model of P-TEFb activation on chromatin (Figure 1).
Figure 1. A model of P-TEFb activation on chromatin by 7SK snRNP chromatin adaptor and P-TEFb release factors.
We envision that each and every major step in our model, including the anchoring of 7SK snRNP to chromatin by Ch-AFs, release of P-TEFb from 7SK snRNP by P-REFs, assembly of P-TEFb with transcription factor and SECs, and sequestration of P-TEFb back into 7SK snRNP upon transcription shutdown, could be regulated. Many questions remain unanswered and interrogating the proposed framework shall yield exciting discoveries in the future.
Matjaz Barboric, Alexandre JC Quaresma, Andrii Bugai
This review was published in Nucleic Acids Research, Oxford University Press. You can read the entire manuscript here:
Ovarian cancer is the 5th most common cause of cancer death in women in United States (6th in Europe), and understanding molecular basis for its genesis is essential for removing the barriers to developing novel prognostic, diagnostic and therapeutic approaches. In high-grade serous ovarian carcinoma, the deadliest form of the disease, the work by The Cancer Genome Atlas (TCGA) consortium has identified CDK12 as one of only nine genes with statistically recurrent somatic mutations. The TCGA finding prompted us to explore whether the mutations in the kinase subunit could be detrimental to the activity and biological function of the novel Cdk12/CycK transcription elongation kinase.
Indeed, we found that most mutations prevented the assembly of the Cdk12/CycK complex and that all mutations except one disabled the kinase activity of Cdk12/CycK in vitro (Figure 1). When we examined positions of the mutations using the published Cdk12/CycK structure, we found that the mutations very likely provoke structural rearrangements detrimental to the Cdk12 activation process. Importantly, our analyses of mRNA expression in patient samples containing the CDK12 mutations revealed that genes essential to the homologous recombination (HR) DNA repair pathway were coordinately downregulated (Figure 1).
When we analyzed these genes in detail, we found that Cdk12/CycK was present at them to promote high Ser2-P levels on the CTD of Pol II, indicating that when active, the Cdk12/CycK complex directly facilitates their expression. Critically, we finally demonstrated that the mutant Cdk12 proteins containing the ovarian carcinoma CDK12 mutations failed to stimulate the error-free DNA double strand break repair by HR (Figure 3).
Together, our study provides the molecular basis of how mutated CDK12 ceases to function in ovarian carcinoma. High-grade serous ovarian carcinoma has been known to possess a highly unstable genome, an enabling characteristic of any cancer. In fact, one half of the tumor cases display genetic and epigenetic defects in the components of the HR repair pathway. Our study demonstrates that mutating CDK12 equips the incipient cancer cells with an alternative source of defects in the HR repair pathway, which is achieved by the collective downregulation of critical HR genes. We propose that CDK12 is a tumor suppressor of which the loss-of-function mutations elicit defects HR-mediated and possibly other DNA repair pathways, leading to genomic instability underlying the genesis of the cancer.
Kingsley M. Ekumi, Hana Paculova, Tina Lenasi, Vendula Pospichalova
We reported our work in Nucleic Acids Research, Oxford University Press. You can read the entire manuscript here: