Biology subjects

Niccum, B. A.

Publications and source records attributed to Niccum, B. A..

6 recordsLinked to original sources

bioRxiv2025

Gastric epithelium from BRCA1 and BRCA2 carriers harbors increased double-stranded DNA damage and augmented growth.

An accumulating body of evidence suggests carriers of a pathogenic germline variant (PGV) in BRCA1 or BRCA2 have increased gastric cancer (GC) risk. BRCA1 and BRCA2 are tumor suppressor genes involved in promoting homologous recombination to repair double-stranded DNA breaks. The aim of this investigation was to identify differences within the gastric epithelium and in patient-derived gastric organoids (PDGOs) between BRCA1 and BRCA2 carriers and non-carriers to determine if evidence of early gastric carcinogenesis exists amongst these carriers. First, using gastric epithelial biopsies, BRCA2 carriers were found to harbor higher expression of the proliferative marker Ki-67 within the antral gastric epithelium and strikingly, biopsies from both BRCA1 and BRCA2 carriers displayed a marked increase in double-stranded DNA damage. These results were further explored using PDGOs, where a growth advantage was observed for both BRCA1 and BRCA2 PDGOs compared to non-carrier PDGOs. Furthermore, both BRCA1 and BRCA2 PDGOs displayed a more pronounced enhancement of Ki-67 expression as well as increased double stranded DNA damage compared to non-carrier PDGOs. Importantly, none of the PDGOs showed signs of BRCA1 or BRCA2 loss of heterozygosity, potentially indicating a haploinsufficient phenotype. Taken together, these novel findings suggest that haploinsufficiency in BRCA1 and BRCA2 carriers may lead to DNA damage in the gastric epithelium, which may serve as an early event contributing to GC development.

cancer biology↗

bioRxiv2021

DNA Replication-transcription conflicts do not significantly contribute to spontaneous mutations due to replication errors in Escherichia coli

Encounters between DNA replication and transcription can cause genomic disruption, particularly when the two meet head-on. Whether these conflicts produce point mutations is debated. This paper presents detailed analyses of a large collection of mutations generated during mutation accumulation experiments with mismatch-repair (MMR) defective Escherichia coli. With MMR absent, mutations are primarily due to DNA replication errors. Overall, there were no differences in the frequencies of base-pair substitutions or small indels (insertion and deletions [≤] 4 bp) in the coding sequences or promoters of genes oriented codirectionally versus head-on to replication. Among a subset of highly expressed genes there was a 2- to 3-fold bias for indels in genes oriented head-on to replication, but this difference was almost entirely due to the asymmetrical genomic locations of tRNA genes containing mononucleotide runs, which are hotspots for indels. No additional orientation bias in mutation frequencies occurred when MMR- strains were also defective for transcription-coupled repair (TCR). However, in contrast to other reports, loss of TCR slightly increased the overall mutation rate, meaning that TCR is antimutagenic. There was no orientation bias in mutation frequencies among the stress-response genes that are regulated by RpoS or induced by DNA damage. Thus, biases in the locations of mutational targets can account for most, if not all, apparent biases in mutation frequencies between genes oriented head-on versus co-directional to replication. In addition, the data revealed a strong correlation of the frequency of base-pair substitutions with gene length, but no correlation with gene expression levels. IMPORTANCEBecause DNA replication and transcription occur on the same DNA template, encounters between the two machines occur frequently. When these encounters are head-to-head, genomic disruption can occur. But, whether replication-transcription conflicts contribute to spontaneous mutations is debated. Analyzing in detail a large collection of mutations generated with mismatch repair defective Escherichia coli strains, we find that across the genome there are no significant differences in mutation frequencies between genes oriented co-directionally versus head-on to replication. Among a subset of highly expressed genes there was a 2- to 3-fold bias for small insertions and deletions in head-on oriented genes, but this difference was almost entirely due to the asymmetrical locations of tRNA genes containing mononucleotide runs, which are hotspots for these mutations. Thus, biases in the positions of mutational target sequences can account for most, if not all, apparent biases in mutation frequencies between genes oriented head-on and co-directionally to replication.

genetics↗

bioRxiv2020

New complexities of SOS-induced untargeted mutagenesis in Escherichia coli as revealed by mutation accumulation and whole-genome sequencing

When its DNA is damaged, Escherichia coli induces the SOS response, which consists of about 40 genes that encode activities to repair or tolerate the damage. Certain alleles of the major SOS-control genes, recA and lexA, cause constitutive expression of the response, resulting in an increase in spontaneous mutations. These mutations, historically called "untargeted", have been the subject of many previous studies. Here we re-examine SOS-induced mutagenesis using mutation accumulation followed by whole-genome sequencing (MA/WGS), which allows a detailed picture of the types of mutations induced as well as their sequence-specificity. Our results confirm previous findings that SOS expression specifically induces transversion base-pair substitutions, with rates averaging about 60-fold above wild-type levels. Surprisingly, the rates of G:C to C:G transversions, normally an extremely rare mutation, were induced an average of 160-fold above wild-type levels. The SOS-induced transversion showed strong sequence specificity, the most extreme of which was the G:C to C:G transversions, 60% of which occurred at the middle base of 5'GGC3'+5'GCC3' sites although these sites represent only 8% of the G:C base pairs in the genome. SOS-induced transversions were also DNA strand biased, occurring, on average, 2- to 4- times more often when the purine was on the leading strand template and the pyrimidine on the lagging strand template than in the opposite orientation. However, the strand bias was also sequence specific, and even of reverse orientation at some sites. By eliminating constraints on the mutations that can be recovered, the MA/WGS protocol revealed new complexities to the nature of SOS "untargeted" mutations. HighlightsO_LIThe SOS response to DNA damage induces "untargeted" mutations C_LIO_LISOS-mutations are revealed by mutation accumulation and whole genome sequencing C_LIO_LISOS-mutations are both sequence and DNA-strand biased C_LIO_LIG:C to C:G transversions are particularly highly induced by SOS C_LIO_LIG:C to C:G transversions are extremely sequence and DNA-strand biased C_LI

genetics↗

bioRxiv2019

Strain-level diversity impacts cheese rind microbiome assembly and function

Taxa that are consistently found across microbial communities are often considered members of a core microbiome. One common assumption is that taxonomically identical core microbiomes will have similar dynamics and functions across communities. However, strain-level genomic and phenotypic variation of core taxa could lead to differences in how core microbiomes assemble and function. Using cheese rinds, we tested whether taxonomically identical core microbiomes isolated from distinct locations have similar assembly dynamics and functional outputs. We first isolated the same three bacterial species (Staphylococcus equorum, Brevibacterium auranticum, and Brachybacterium alimentarium) from nine cheeses produced in different regions of the United States and Europe. Comparative genomics identified distinct phylogenetic clusters and significant variation in genome content across the nine core microbiomes. When we assembled each core microbiome with initially identical compositions, community structure diverged over time resulting in communities with different dominant taxa. The core microbiomes had variable responses to abiotic (high salt) and biotic (the fungus Penicillium) perturbations, with some communities showing no response and others substantially shifting in composition. Functional differences were also observed across the nine core communities, with considerable variation in pigment production (light yellow to orange) and composition of volatile organic compound profiles emitted from the rinds (nutty to sulfury). Our work demonstrates that core microbiomes isolated from independent communities may not function in the same manner due to strain-level variation of core taxa. Strain-level diversity across core cheese rind microbiomes may contribute to variability in the aesthetics and quality of surface-ripened cheeses.

microbiology↗

bioRxiv2019

The symmetrical pattern of base-pair substitutions rates across the chromosome in Escherichia coli has multiple causes

Mutation accumulation experiments followed by whole-genome sequencing have revealed that for several bacterial species the rate of base-pair substitutions is not constant across the chromosome but varies in a wave-like pattern symmetrical about the origin of replication. The experiments reported here demonstrate that in Escherichia coli several interacting factors determine the wave. Perturbing replication timing, progression, or the structure of the terminus disrupts the pattern. Biases in error-correction by proofreading and mismatch repair are major factors. The activities of the nucleoid binding proteins, HU and Fis, are important, suggesting that mutation rates increase when highly structured DNA is replicated. These factors should apply to most bacterial, and possibly eukaryotic, genomes, and imply that different areas of the genome evolve at different rates.

microbiology↗

bioRxiv2018

Determinants of base-pair substitution patterns revealed by whole-genome sequencing of DNA mismatch repair defective Escherichia coli

Mismatch repair (MMR) is a major contributor to replication fidelity, but its impact varies with sequence context and the nature of the mismatch. Mutation accumulation experiments followed by whole-genome sequencing of MMR-defective E. coli strains yielded {approx}30,000 base-pair substitutions, revealing mutational patterns across the entire chromosome. The base-pair substitution spectrum was dominated by A:T > G:C transitions, which occurred predominantly at the center base of 5'NAC3'+5'GTN3' triplets. Surprisingly, growth on minimal medium or at low temperature attenuated these mutations. Mononucleotide runs were also hotspots for base-pair substitutions, and the rate at which these occurred increased with run length. Comparison with {approx}2000 base-pair substitutions accumulated in MMR-proficient strains revealed that both kinds of hotspots appeared in the wild-type spectrum and so are likely to be sites of frequent replication errors. In MMR-defective strains transitions were strand biased, occurring twice as often when A and C rather than T and G were on the lagging-strand template. Loss of nucleotide diphosphate kinase increases the cellular concentration of dCTP, which resulted in increased rates of mutations due to misinsertion of C opposite A and T. In an mmr ndk double mutant strain, these mutations were more frequent when the template A and T were on the leading strand, suggesting that lagging-strand synthesis was more error-prone or less well corrected by proofreading than was leading strand synthesis.

genetics↗