Histone H2AX Y142 phosphorylation is a low abundance modification

We employ targeted mass spectrometry to compare the levels of H2AX S139 phosphorylation (γH2AX) and Y142 phosphorylation. We use synthetic peptides to facilitate MS optimisation and estimate relative detection efficiencies for the different modifications. Despite phosphopeptide enrichment from large amounts of starting material, we are unable to detect endogenous H2AX Y142 phosphorylation, indicating that it is present in low abundance (<1%). We also calculate the relative levels of H2AX compared to other H2A isoforms and quantify the proportion of H2AX that is phosphorylated on S139 (γH2AX) after ionising radiation

Multi-omic profiling reveals the ataxia protein sacsin is required for integrin trafficking and synaptic organization

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a childhood-onset cerebellar ataxia caused by mutations in SACS, which encodes the protein sacsin. Cellular ARSACS phenotypes include mitochondrial dysfunction, intermediate filament disorganization, and progressive death of cerebellar Purkinje neurons. It is unclear why the loss of sacsin causes these deficits or why they manifest as cerebellar ataxia. Here, we perform multi-omic profiling in sacsin knockout (KO) cells and identify alterations in microtubule dynamics and mislocalization of focal adhesion (FA) proteins, including multiple integrins. Deficits in FA structure, signaling, and function can be rescued by targeting PTEN, a negative regulator of FA signaling. ARSACS mice possess mislocalization of ITGA1 in Purkinje neurons and synaptic disorganization in the deep cerebellar nucleus (DCN). The sacsin interactome reveals that sacsin regulates interactions between cytoskeletal and synaptic adhesion proteins. Our findings suggest that disrupted trafficking of synaptic adhesion proteins is a causal molecular deficit in ARSACS

Multi-omic profiling reveals the ataxia protein sacsin is required for integrin trafficking and synaptic organization.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a childhood-onset cerebellar ataxia caused by mutations in SACS, which encodes the protein sacsin. Cellular ARSACS phenotypes include mitochondrial dysfunction, intermediate filament disorganization, and progressive death of cerebellar Purkinje neurons. It is unclear why the loss of sacsin causes these deficits or why they manifest as cerebellar ataxia. Here, we perform multi-omic profiling in sacsin knockout (KO) cells and identify alterations in microtubule dynamics and mislocalization of focal adhesion (FA) proteins, including multiple integrins. Deficits in FA structure, signaling, and function can be rescued by targeting PTEN, a negative regulator of FA signaling. ARSACS mice possess mislocalization of ITGA1 in Purkinje neurons and synaptic disorganization in the deep cerebellar nucleus (DCN). The sacsin interactome reveals that sacsin regulates interactions between cytoskeletal and synaptic adhesion proteins. Our findings suggest that disrupted trafficking of synaptic adhesion proteins is a causal molecular deficit in ARSACS

The amyloid architecture provides a scaffold for enzyme-like catalysts

Natural biological enzymes possess catalytic sites that are generally surrounded by a large three-dimensional scaffold. However, the proportion of the protein molecule that participates in the catalytic reaction is relatively small. The generation of artificial or miniature enzymes has long been a focus of research because enzyme mimetics can be produced with high activity at low cost. These enzymes aim to mimic the active sites without the additional architecture contributed by the protein chain. Previous work has shown that amyloidogenic peptides are able to self-assemble to create an active site that is capable of binding zinc and catalysing an esterase reaction. Here, we describe the structural characterisation of a set of designed peptides that form an amyloid-like architecture and reveal that their capability to mimic carbonic anhydrase and serve as enzyme-like catalysts is related to their ability to self-assemble. These amyloid fibril structures can bind the metal ion Zn2+ via a three-dimensional arrangement of His residues created by the amyloid architecture. Our results suggest that the catalytic efficiency of amyloid-like assembly is not only zinc-dependent but also depends on an active centre created by the peptides which is, in turn, dependent on the ordered architecture. These fibrils have good esterase activity, and they may serve as good models for the evolution of modern-day enzymes. Furthermore, they may be useful in designing self-assembling fibrils for applications as metal ion catalysts. This study also demonstrates that the ligands surrounding the catalytic site affect the affinity of the zinc-binding site to bind the substrate contributing to the enzymatic activity of the assembled peptides

250 years of hybridisation between two biennial herb species without speciation.

Hybridization between plant species can generate novel morphological diversity and lead to speciation at homoploid or polyploid levels. Hybrids between biennial herbs Tragopogon pratensis and T. porrifolius have been studied in experimental and natural populations for over 250 years. Here we examine their current status in natural populations in southeast England. All hybrids found were diploid; they tended to grow taller and with more buds than their parental species; many showed partial fertility; a few showed evidence of backcrossing. However, we found no evidence to suggest that the hybrids are establishing as a new species, nor can we find literature documenting speciation of these hybrids elsewhere. This lack of speciation despite at least 250 years of hybridization contrasts with the fact that both parental species have formed new allopolyploid species through hybridization with another diploid, T. dubius. Understanding why hybrids often do not speciate, despite repeated opportunities, would enhance our understanding of both the evolutionary process and risk assessments of invasive species