DNA polymerase λ promotes error-free replication through Watson-Crick impairing N1-methyl-deoxyadenosine adduct in conjunction with DNA polymerase ζ.

In a previous study, we showed that replication through the N1-methyl-deoxyadenosine (1-MeA) adduct in human cells is mediated via three different Polι/Polθ, Polη, and Polζ-dependent pathways. Based on biochemical studies with these Pols, in the Polι/Polθ pathway, we inferred a role for Polι in the insertion of a nucleotide (nt) opposite 1-MeA and of Polθ in extension of synthesis from the inserted nt; in the Polη pathway, we inferred that this Pol alone would replicate through 1-MeA; in the Polζ pathway, however, the Pol required for inserting an nt opposite 1-MeA had remained unidentified. In this study, we provide biochemical and genetic evidence for a role for Polλ in inserting the correct nt T opposite 1-MeA, from which Polζ would extend synthesis.

Bi-allelic MCM10 variants associated with immune dysfunction and cardiomyopathy cause telomere shortening.

Minichromosome maintenance protein 10 (MCM10) is essential for eukaryotic DNA replication. Here, we describe compound heterozygous MCM10 variants in patients with distinctive, but overlapping, clinical phenotypes: natural killer (NK) cell deficiency (NKD) and restrictive cardiomyopathy (RCM) with hypoplasia of the spleen and thymus. To understand the mechanism of MCM10-associated disease, we modeled these variants in human cell lines.

A novel role of DNA polymerase λ in translesion synthesis in conjunction with DNA polymerase ζ.

By extending synthesis opposite from a diverse array of DNA lesions, DNA polymerase (Pol) ζ performs a crucial role in translesion synthesis (TLS). In yeast and cancer cells, Rev1 functions as an indispensable scaffolding component of Polζ and it imposes highly error-prone TLS upon Polζ. However, for TLS that occurs during replication in normal human cells, Rev1 functions instead as a scaffolding component of Pols η, ι, and κ and Rev1-dependent TLS by these Pols operates in a predominantly error-free manner.

Error-Prone Replication through UV Lesions by DNA Polymerase θ Protects against Skin Cancers.

Cancers from sun-exposed skin accumulate "driver" mutations, causally implicated in oncogenesis. Because errors incorporated during translesion synthesis (TLS) opposite UV lesions would generate these mutations, TLS mechanisms are presumed to underlie cancer development. To address the role of TLS in skin cancer formation, we determined which DNA polymerase is responsible for generating UV mutations, analyzed the relative contributions of error-free TLS by Polη and error-prone TLS by Polθ to the replication of UV-damaged DNA and to genome stability, and examined the incidence of UV-induced skin cancers in Polθ, Polη, and Polθ Polη mice.

Identification of two functional PCNA-binding domains in human DNA polymerase κ.

Previously, we have shown that human DNA polymerase (Pol) η has two functional PCNA-binding motifs, PIP1 and PIP2, and that a C-terminal deletion of Polη that lacks the ubiquitin-binding UBZ domain and the PIP2 domain but retains the PIP1 domain promotes normal levels of translesion synthesis (TLS) opposite a cis-syn TT dimer in human cells. Here, we identify two PIP domains in Polκ and show that TLS occurs normally in human fibroblast cells in which the pip1 or pip2 mutant Polκ is expressed, but mutational inactivation of both PIP domains renders Polκ nonfunctional in TLS opposite the thymine glycol lesion. Thus, the two PIP domains of Polκ function redundantly in TLS opposite this DNA lesion in human cells.

DNA clasping by mycobacterial HU: the C-terminal region of HupB mediates increased specificity of DNA binding.

Background: HU a small, basic, histone like protein is a major component of the bacterial nucleoid. E. coli has two subunits of HU coded by hupA and hupB genes whereas Mycobacterium tuberculosis (Mtb) has only one subunit of HU coded by ORF Rv2986c (hupB gene).

A novel nucleoid-associated protein of Mycobacterium tuberculosis is a sequence homolog of GroEL.

The Mycobacterium tuberculosis genome sequence reveals remarkable absence of many nucleoid-associated proteins (NAPs), such as HNS, Hfq or DPS. In order to characterize the nucleoids of M. tuberculosis, we have attempted to identify NAPs, and report an interesting finding that a chaperonin-homolog, GroEL1, is nucleoid associated.