Zenker, J. | Search bioRxiv

bioRxiv2023

Epsilon tubulin is an essential determinant of microtubule-based structures in male germ cells

Alpha, beta and gamma tubulins are essential building blocks for all eukaryotic cells. The functions of the non-canonical tubulins, delta, epsilon and zeta, however, remain poorly understood and their requirement in mammalian development untested. Herein we have used a spermatogenesis model to define epsilon tubulin (TUBE1) function in mice. We show that TUBE1 is essential for the function of multiple complex microtubule arrays, including the meiotic spindle, axoneme and manchette and in its absence, there is a dramatic loss of germ cells and male sterility. Through examining axoneme structure, we identify differences in TUBE1 function between somatic and germ cells and potentially between species. Moreover, we provide evidence for the interplay between TUBE1 and katanin-mediated microtubule severing, and for the sub-specialization of individual katanin paralogs in the regulation of specific microtubule arrays.

developmental biology↗

bioRxiv2022

Systematic loss of function screens identify pathway specific functional circular RNAs

Circular RNAs (circRNAs) are covalently closed single stranded RNAs that are produced by RNA back-splicing. A small number of circRNAs have been implicated as functional, however, we still lack systematic understanding of cellular processes and signalling pathways that are regulated by circRNAs. A major gap in understanding circRNA function is the ability to define pathways that are regulated by circRNAs. Here, we generated a pooled shRNA library targeting the back-splice junction of 3,354 human circRNAs that are expressed at low to high levels in humans. We used this library for loss of function proliferation screens in a panel of 18 cancer cell lines from four tissue types that harbour mutations leading to constitutive activity of defined pathways. Using this dataset, we identify context specific and non-specific circRNAs. We validated these observations with a secondary screen and uncovered a role for circRERE, a cell essential circRNA that regulates ferroptosis. Furthermore, we characterised the functional roles of pathway-specific circRNA, circSMAD2, a novel regulator of the WNT pathway and circMTO1, a regulator of MAPK signalling in a PTEN dependent manner. Our work sheds light on molecular pathways regulated by circRNAs and provides a catalogue of circRNAs with a measurable function in human cells.

genetics↗