Biomedical subjects

Ekaterina V Mirkin

Publications and source records attributed to Ekaterina V Mirkin.

3 recordsLinked to original sources

PubMed2006

Transcription regulatory elements are punctuation marks for DNA replication.

Collisions between DNA replication and transcription significantly affect genome organization, regulation, and stability. Previous studies have described collisions between replication forks and elongating RNA polymerases. Although replication collisions with the transcription-initiation or -termination complexes are potentially even more important because most genes are not actively transcribed during DNA replication, their existence and mechanisms remained unproven. To address this matter, we have designed a bacterial promoter that binds RNA polymerase and maintains it in the initiating mode by precluding the transition into the elongation mode. By using electrophoretic analysis of replication intermediates, we have found that this steadfast transcription-initiation complex inhibits replication fork progression in an orientation-dependent manner during head-on collisions. Transcription terminators also appeared to attenuate DNA replication, but in the opposite, codirectional orientation. Thus, transcription regulatory signals may serve as "punctuation marks" for DNA replication in vivo.

AT Rich Sequence↗

PubMed2005

Mechanisms of transcription-replication collisions in bacteria.

While collisions between replication and transcription in bacteria are deemed inevitable, the fine details of the interplay between the two machineries are poorly understood. In this study, we evaluate the effects of transcription on the replication fork progression in vivo, by using electrophoresis analysis of replication intermediates. Studying Escherichia coli plasmids, which carry constitutive or inducible promoters in different orientations relative to the replication origin, we show that the mutual orientation of the two processes determines their mode of interaction. Replication elongation appears not to be affected by transcription proceeding in the codirectional orientation. Head-on transcription, by contrast, leads to severe inhibition of the replication fork progression. Furthermore, we evaluate the mechanism of this inhibition by limiting the area of direct contact between the two machineries. We observe that replication pausing zones coincide exactly with transcribed DNA segments. We conclude, therefore, that the replication fork is most likely attenuated upon direct physical interaction with the head-on transcription machinery.

DNA Replication↗

PubMed2003

Schizosaccharomyces pombe Ddb1 is functionally linked to the replication checkpoint pathway.

Schizosaccharomyces pombe Ddb1 is homologous to the mammalian DDB1 protein, which has been implicated in damaged-DNA recognition and global genomic repair. However, a recent study suggested that the S. pombe Ddb1 is involved in cell division and chromosomal segregation. Here, we provide evidence that the S. pombe Ddb1 is functionally linked to the replication checkpoint control gene cds1. We show that the S. pombe strain lacking ddb1 has slow growth due to delayed replication progression. Flow cytometric analysis shows an extensive heterogeneity in DNA content. Furthermore, the Deltaddb1 strain is hypersensitive to UV irradiation in S phase and is unable to tolerate a prolonged replication block imposed by hydroxyurea. Interestingly, the Deltaddb1 strain exhibits a high level of the Cds1 kinase activity during passage through S phase. Moreover, mutation of the cds1 gene relieves the defects observed in Deltaddb1 strain. The results suggest that many of the defects observed in Deltaddb1 cells are linked to an aberrant activation of Cds1, and that Ddb1 is functionally linked to Cds1.

Checkpoint Kinase 2↗