Biology subjects

Sivakumar, M.

Publications and source records attributed to Sivakumar, M..

3 recordsLinked to original sources

bioRxiv2026

16S rRNA Modifications Are Dispensable for Viability but Collectively Optimize Ribosome Biogenesis and Translation Initiation in Escherichia coli

Ribosomal RNAs contain numerous conserved nucleotide modifications, yet the functional importance of most of these modifications remains unclear. In Escherichia coli, deletion of individual 16S rRNA modification enzymes generally produces only minor phenotypes, raising questions about their biological significance. Here, we generated a comprehensive collection of deletion mutants lacking individual and combined 16S rRNA modifications, culminating in a strain lacking all known 30S ribosomal subunit modifications. Despite the absence of all known 16S rRNA modifications, cells remained viable, exhibiting a fitness defect of ~30% at 37 {degrees}C that increased to ~50% at 20 {degrees}C, consistent with impaired ribosome biogenesis. We identified strong epistatic interactions between modifications in the 3' major and 3' minor domains of 16S rRNA, resulting in disproportionately large effects on both fitness and antibiotic susceptibility. Live-cell single-molecule tracking revealed a marked increase in the fraction of non-translating ribosomes and a prolonged time required to enter productive translation, whereas translational elongation by actively engaged 70S ribosomes remained largely unaffected. In addition, fluorescence-based measurements showed that unmodified ribosomes exhibited increased stringency during translation initiation, reducing utilization of near-cognate start codons. Together, these findings demonstrate that 16S rRNA modifications are not essential for viability but collectively enhance the efficiency, robustness, and fidelity of ribosome assembly and translational initiation.

microbiology↗

bioRxiv2023

Phenotyping of lymphoproliferative tumours generated in xenografts of non-small cell lung cancer

Patient-derived xenograft (PDX) models involve the engraftment of tumour tissue in immunocompromised mice and represent an important pre-clinixtcal oncology research. A limitation of non-small cell lung cancer (NSCLC) PDX model derivation in NOD-scid IL2Rgammanull (NSG) mice is that a subset of initial engraftments are of lymphocytic, rather than tumour origin. In the lung TRACERx PDX pipeline, lymphoproliferations occurred in 17.8% of lung adenocarcinoma and 10% of lung squamous cell carcinoma transplantations, despite none of these patients having a prior or subsequent clinical history of lymphoproliferative disease. Lymphoproliferations were predominantly human CD20+ B cells and had the immunophenotype expected for post-transplantation diffuse large B cell lymphoma. All lymphoproliferations expressed Epstein-Barr-encoded RNAs (EBER). Analysis of immunoglobulin light chain gene rearrangements in three tumours where multiple tumour regions had resulted in lymphoproliferations suggested that each had independent clonal origins. Overall, these data suggest the presence of B cell clones with lymphoproliferative potential within primary NSCLC tumours that are under continuous immune surveillance. Since these cells can be expanded following transplantation into NSG mice, our data highlight the value of quality control measures to identify lymphoproliferations within xenograft pipelines and support the incorporation of strategies to minimise lymphoproliferations during the early stages of xenograft establishment pipelines. To present the histology data herein, we developed a Python-based tool for generating patient-level pathology overview figures from whole-slide image files; PATHOverview is available on GitHub (https://github.com/EpiCENTR-Lab/PATHOverview).

cancer biology↗

bioRxiv2023

Genomic evolution of non-small cell lung cancer patient-derived xenograft models

Patient-derived xenograft (PDX) models of cancer, developed through injection of patient tumour cells into immunocompromised mice, have been widely adopted in preclinical studies, as well as in precision oncology approaches. However, the extent to which PDX models represent the underlying genetic diversity of a patients tumour and the extent of on-going genomic evolution in PDX models are incompletely understood, particularly in the context of heterogeneous cancers such as non-small cell lung cancer (NSCLC). To investigate the depiction of intratumour heterogeneity by PDX models, we derived 47 new subcutaneous multi-region PDX models from 22 patients with primary NSCLC enrolled in the clinical longitudinal cohort study TRACERx. By analysing whole exome sequencing data from primary tumours and PDX models, we find that PDX establishment creates a genomic bottleneck, with 76% of PDX models being derived from a single primary tumour subclone. Despite this, multiple primary tumour subclones were capable of PDX establishment in regional PDX models, indicating that PDX libraries derived from multiple tumour regions can capture intratumour heterogeneity. Acquisition of somatic mutations continued during PDX model expansion, and was associated with APOBEC- or mismatch repair deficiency-induced mutational signatures in a subset of models. Overall, while NSCLC PDX models retain truncal genomic alterations, the absence of subclonal heterogeneity representative of the primary tumour is a major limitation. Our results emphasise the importance of characterising and monitoring intratumour heterogeneity in the context of pre-clinical cancer studies.

cancer biology↗