Nibrin Forkhead-associated Domain and Breast Cancer C-terminal Domain Are Both Required for Nuclear Focus Formation and Phosphorylation

HighlightsPeptidyl arginine deiminase 2 (PADI2) citrullinates arginine1810 (cit1810) present at carboxy-terminal domain of RNA polymerase II (RNAP2-CTD).PADI2 and R1810 of RNAP2-CTD regulate transcription and proliferation of breast cancer cells.Absence of cit1810 at RNAP2-CTD leads to RNAP2 accumulation at proximal promoter regions.Cit1810 at RNAP2-CTD facilitate interaction with P-TFEb complex.SUMMARYThe post-translational modification of key residues at the carboxy-terminal domain of RNA polymerase II (RNAP2-CTD), coordinates transcription, splicing, and RNA processing by modulating its capacity to act as a landing platform for a variety of protein complexes. Here, we identify a new modification at the CTD, the deimination of arginine and its conversion to citrulline by peptidyl arginine deiminase 2 (PADI2), an enzyme that has been associated with several diseases including cancer. We show that among PADI family members, only PADI2 citrullinates R1810 (Cit1810) at repeat 31 of the CTD. Depletion of PADI2 or loss of R1810 result in accumulation of RNAP2 at transcription start sites, reduced gene expression and inhibition of cell proliferation. Cit1810 is needed for interaction with the P-TEFb (positive transcription elongation factor b) kinase complex and for its recruitment to chromatin. In this way, CTD-Cit1810 favors RNAP2 pause release and efficient transcription in breast cancer cells.

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Rec2 Interplay with both Brh2 and Rad51 Balances Recombinational Repair in Ustilago maydis

Molecular and Cellular Biology ◽

10.1128/mcb.26.2.678-688.2006 ◽

2006 ◽

Vol 26 (2) ◽

pp. 678-688 ◽

Cited By ~ 17

Author(s):

Milorad Kojic ◽

Qingwen Zhou ◽

Michael Lisby ◽

William K. Holloman

Keyword(s):

High Frequency ◽

Ustilago Maydis ◽

Radiation Sensitivity ◽

Recombinational Repair ◽

Focus Formation ◽

Transgenic Expression ◽

Rad51 Focus ◽

Nuclear Focus ◽

Extreme Degree ◽

Radiation Induced

ABSTRACT Rec2 is the single Rad51 paralog in Ustilago maydis. Here, we find that Rec2 is required for radiation-induced Rad51 nuclear focus formation but that Rec2 foci form independently of Rad51 and Brh2. Brh2 foci also form in the absence of Rad51 and Rec2. By coprecipitation from cleared extracts prepared from Escherichia coli cells expressing the proteins, we found that Rec2 interacts physically not only with Rad51 and itself but also with Brh2. Transgenic expression of Brh2 in rec2 mutants can effectively restore radiation resistance, but the frequencies of spontaneous Rad51 focus formation and allelic recombination are elevated. The Dss1-independent Brh2-RPA70 fusion protein is also active in restoring radiation sensitivity of rec2 but is hyperactive to an extreme degree in allelic recombination and in suppressing the meiotic block of rec2. However, the high frequency of chromosome missegregation in meiotic products is an indicator of a corrupted process. The results demonstrate that the importance of Rec2 function is not only in stimulating recombination activity but also in ensuring that recombination is properly controlled.

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DNA Cross-Link Repair Protein SNM1A Interacts with PIAS1 in Nuclear Focus Formation

Molecular and Cellular Biology ◽

10.1128/mcb.24.24.10733-10741.2004 ◽

2004 ◽

Vol 24 (24) ◽

pp. 10733-10741 ◽

Cited By ~ 55

Author(s):

Masamichi Ishiai ◽

Masayo Kimura ◽

Keiko Namikoshi ◽

Mitsuyoshi Yamazoe ◽

Kazuhiko Yamamoto ◽

...

Keyword(s):

Dna Repair ◽

Point Mutations ◽

Translesion Synthesis ◽

X Rays ◽

Cross Link ◽

Focus Formation ◽

Nuclear Focus ◽

Repair Protein ◽

Repair Pathways ◽

Nuclear Foci

ABSTRACT The yeast SNM1/PSO2 gene specifically functions in DNA interstrand cross-link (ICL) repair, and its role has been suggested to be separate from other DNA repair pathways. In vertebrates, there are three homologs of SNM1 (SNM1A, SNM1B, and SNM1C/Artemis; SNM1 family proteins) whose functions are largely unknown. We disrupted each of the SNM1 family genes in the chicken B-cell line DT40. Both SNM1A- and SNM1B-deficient cells were sensitive to cisplatin but not to X-rays, whereas SNM1C/Artemis-deficient cells exhibited sensitivity to X-rays but not to cisplatin. SNM1A was nonepistatic with XRCC3 (homologous recombination), RAD18 (translesion synthesis), FANCC (Fanconi anemia), and SNM1B in ICL repair. SNM1A protein formed punctate nuclear foci depending on the conserved SNM1 (metallo-β-lactamase) domain. PIAS1 was found to physically interact with SNM1A, and they colocalized at nuclear foci. Point mutations in the SNM1 domain, which disrupted the interaction with PIAS1, led to mislocalization of SNM1A in the nucleus and loss of complementation of snm1a cells. These results suggest that interaction between SNM1A and PIAS1 is required for ICL repair.

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