Biology subjects

Venkatesh, N.

Publications and source records attributed to Venkatesh, N..

3 recordsLinked to original sources

bioRxiv2026

Exclusion from DNA double-strand breaks is a pervasive feature of the hnRNP family

Heterogeneous nuclear ribonucleoproteins (hnRNPs) are abundant RNA-binding proteins that have been linked individually to the DNA double-strand break (DSB) response, but their behaviour at damaged chromatin has only been examined one protein at a time and under disparate conditions. Here, we compare the core hnRNP family side by side by laser microirradiation and live-cell imaging. We find that 13 of 14 members are excluded from damaged chromatin, 10 directly and 3 after a transient phase of recruitment, establishing exclusion rather than accumulation as the predominant behaviour of this family. We characterised this behaviour in detail for hnRNPH1, which has not previously been linked directly to the DSB response. Its exclusion is sustained for several hours and also occurs at nuclease-induced breaks at a defined genomic locus. Importantly, exclusion is independent of the signalling pathways that drive protein recruitment to breaks and of transcriptional elongation, and instead requires the GYR low-complexity domain, with RNA binding contributing redundantly through three quasi-RNA recognition motifs. Finally, tethering hnRNPH1 to breaks through the RNF8 FHA domain selectively impairs 53BP1 accumulation without affecting {gamma}-H2AX, MDC1 or RPA, and hnRNPH1 is excluded from 53BP1 nuclear bodies in unchallenged cells. Together, these findings establish protein exclusion as an active component of the DSB response and show that the presence of an hnRNP at damaged chromatin can determine which repair factors assemble at DNA break sites.

cell biology↗

bioRxiv2023

Ataxin-2 polyglutamine expansions aberrantly sequester TDP-43, drive ribonucleoprotein condensate transport dysfunction and suppress local translation

Altered RNA metabolism is a common pathogenic mechanism linked to familial and sporadic Amyotrophic lateral sclerosis (ALS). ALS is characterized by mislocalization and aggregation of TDP-43, an RNA-binding protein (RBP) with multiple roles in post-transcriptional RNA processing. Recent studies have identified genetic interactions between TDP-43 and Ataxin-2, a polyglutamine (polyQ) RBP in which intermediate length polyQ expansions confer increased ALS risk. Here, we used live-cell confocal imaging, photobleaching and translation reporter assays to study the localization, transport dynamics and mRNA regulatory functions of TDP-43/Ataxin-2 in rodent primary cortical neurons. We show that Ataxin-2 polyQ expansions aberrantly sequester TDP-43 within ribonucleoprotein (RNP) condensates, and disrupt both its motility along the axon and liquid-like properties. Our data suggest that Ataxin-2 governs motility and translation of neuronal RNP condensates and that Ataxin-2 polyQ expansions fundamentally perturb spatial localization of mRNA and suppress local translation. Overall, these results indicate Ataxin-2 polyQ expansions have detrimental effects on stability, localization, and translation of transcripts critical for axonal and cytoskeletal integrity, particularly important for motor neurons.

cell biology↗

bioRxiv2020

Student-focused development of a next-generation centrifuge force microscope

Advanced biological molecule force probing methods such as atomic force microscopy and optical tweezers used to quantify forces at the single-molecule level are expensive and require extensive training and technical knowledge. However, the technologies underlying a centrifuge force microscope (CFM) are relatively straight forward, allowing for construction by labs with relatively low budgets and minimal training. Design ideas from previously constructed CFMs served as a guide in the development of this CFM. There were two primary goals: first, to develop an inexpensive, functional CFM using off-the-shelf and 3D printed parts; and second, to do so in the context of providing an educational experience for a broad range of students. The team included high school students and undergraduates from local high schools, the University of Minnesota, and other local higher education institutions. This project created an environment for student-focused development of the CFM that fostered active learning, individual ownership, as well as excellence in research. The instrument discussed herein represents a fully functional CFM designed and built by a postdoctoral researcher and a graduate student who together mentored several high school and undergraduate students. STATEMENT OF SIGNIFICANCEThe presented centrifuge force microscope (CFM) builds on features of existing designs specifically engineered for probing macromolecular force interactions at the single-molecule level. In the coming years, more versatile and modular CFM designs will be utilized in the force spectroscopy field, and the presented design is a step in that direction. In addition to constructing a functional instrument, true student ownership of the project design was equally an end goal. Students from high school through graduate school were included, and the project was structured so that everyone was seen as peers. This active learning project allowed students to acquire scientific concepts and techniques and apply them to real-life situations.

biophysics↗