Biology subjects

Maudlin, I.

Publications and source records attributed to Maudlin, I..

3 recordsLinked to original sources

bioRxiv2026

Inhibition of CDK7, CDK9 or CDK12 affects phosphorylation of human SPT5.

SPT5, complexed with SPT4 in the DRB-sensitivity-inducing factor, DSIF, is a critical regulator of transcription elongation that acts negatively in the unphosphorylated form and positively when phosphorylated. In vivo, phosphorylation of multiple sites in the largely unstructured linker and C-terminal regions of the protein facilitates release from an early elongation checkpoint and processive transcription elongation. Transcription elongation factors, including the PAF complex (PAF1C), interact with phosphorylated SPT5 and phosphorylation of residues S666 and T806 are associated with the positive elongation functions of SPT5. Here, using in vitro phosphorylation of regions of the SPT5 C-terminus followed by GST-mediated pull-down we confirm that the RTF1 subunit of PAF1C interacts with residues 750-850 only when phosphorylated. Antibodies we raised to phospho-S666 or phospho-T806 are specific for these sites phosphorylated in vitro and in vivo. Western analysis of endogenous SPT5 in HEK293 cells using these antibodies before and after inhibition of analog-sensitive versions of CDK7, CDK9 and CDK12 indicate that all three kinases play roles in the phosphorylation of SPT5, although inhibition of CDK9 has the most marked effect. CDK9 inhibition also inhibits the association of PAF1C subunits with SPT5 in vivo and affects phosphorylation of chromatin-associated SPT5 as measured by chromatin immunoprecipitation (ChIP).

molecular biology↗

bioRxiv2026

A genome-wide atlas of meiotic recombination intermediates reveals distinct modes of DNA repair that direct crossovers away from transcriptionally marked genes

Crossovers are essential for accurate chromosome segregation in meiosis. Yet the programmed DNA double-strand breaks that initiate them frequently occur in genes and pose a risk to transcription required for gametogenesis. How meiotic cells reconcile these competing demands has remained unclear. Here, we generate a genome-wide in vivo atlas of meiotic recombination intermediates across [~]42,000 hotspots by mapping repair proteins BLM, HFM1, and RPA in wild-type and genome-engineered mutant mouse testes. These maps reveal two distinct modes of break repair: a fast-resolving class with short-lived intermediates that are repaired predominantly as non-crossovers, and a slower class with persistent intermediates that give rise to nearly all crossovers. Fast-resolving hotspots occur almost exclusively within a deeply conserved set of [~]4,500 genes marked by structural and chromatin features established during an early stage of meiotic transcription. This transcriptional memory predicts repair fate with high accuracy across mouse subspecies and sexes. Across widely diverged mammals, including humans and cattle, orthologous genes show similar crossover suppression. Our findings reveal an early bifurcation between crossover and non-crossover repair that is governed by the transcriptional context of meiotic breaks. Together, they establish an evolutionarily conserved principle in which crossovers are directed away from transcriptionally important genes, thereby safeguarding gene function and shaping their evolution.

genomics↗

bioRxiv2021

Black-necked spitting cobra (Naja nigricollis) phospholipases A2 cause Trypanosoma brucei death by blocking endocytosis through the flagellar pocket

African trypanosomes, such as Trypanosoma brucei, are flagellated protozoa which proliferate in mammals and cause a variety of diseases in people and animals. In a mammalian host, the external face of the African trypanosome plasma membrane is covered by a densely packed coat formed of variant surface glycoprotein (VSG), which counteracts the host adaptive immune response by antigenic variation. The VSG is attached to the external face of the plasma membrane by covalent attachment of the C-terminus to a glycosylphosphatidylinositol. As the trypanosome grows, newly synthesised VSG is added to the plasma membrane by vesicle fusion to the flagellar pocket, the sole location of exo- and endocytosis. Snake venoms contain dozens of components including proteases and phospholipases. Here, we investigated the effect of Naja nigricollis on T. brucei with the aim of describing the response of the trypanosome to hydrolytic attack on the VSG. We found no evidence for VGS hydrolysis however N. nigricollis venom caused: (i) an enlargement of the flagellar pocket, (ii) the Rab11 positive endosomal compartments to adopt an abnormal dispersed localisation, and (iii) a cell cycle arrest prior to cytokinesis. A single protein family, the phospholipases A2s present in N. nigricollis venom, was necessary and sufficient for the effects. This study provides new molecular insight into T. brucei biology and possibly describes mechanisms that could be exploited for T. brucei targeting.

microbiology↗