Deep proteomics identifies shared molecular pathway alterations in synapses of patients with schizophrenia and bipolar disorder and mouse model.

Synaptic dysfunction is implicated in the pathophysiology of schizophrenia (SCZ) and bipolar disorder (BP). We use quantitative mass spectrometry to carry out deep, unbiased proteomic profiling of synapses purified from the dorsolateral prefrontal cortex of 35 cases of SCZ, 35 cases of BP, and 35 controls. Compared with controls, SCZ and BP synapses show substantial and similar proteomic alterations. Network analyses reveal upregulation of proteins associated with autophagy and certain vesicle transport pathways and downregulation of proteins related to synaptic, mitochondrial, and ribosomal function in the synapses of individuals with SCZ or BP. Some of the same pathways are similarly dysregulated in the synaptic proteome of mutant mice deficient in Akap11, a recently discovered shared risk gene for SCZ and BP. Our work provides biological insights into molecular dysfunction at the synapse in SCZ and BP and serves as a resource for understanding the pathophysiology of these disorders.

Brain-region-specific changes in neurons and glia and dysregulation of dopamine signaling in Grin2a mutant mice.

A genetically valid animal model could transform our understanding of schizophrenia (SCZ) disease mechanisms. Rare heterozygous loss-of-function (LoF) mutations in GRIN2A, encoding a subunit of the NMDA receptor, greatly increase the risk of SCZ. By transcriptomic, proteomic, and behavioral analyses, we report that heterozygous Grin2a mutant mice show (1) large-scale gene expression changes across multiple brain regions and in neuronal (excitatory and inhibitory) and non-neuronal cells (astrocytes and oligodendrocytes), (2) evidence of hypoactivity in the prefrontal cortex (PFC) and hyperactivity in the hippocampus and striatum, (3) an elevated dopamine signaling in the striatum and hypersensitivity to amphetamine-induced hyperlocomotion (AIH), (4) altered cholesterol biosynthesis in astrocytes, (5) a reduction in glutamatergic receptor signaling proteins in the synapse, and (6) an aberrant locomotor pattern opposite of that induced by antipsychotic drugs. These findings reveal potential pathophysiologic mechanisms, provide support for both the "hypo-glutamate" and "hyper-dopamine" hypotheses of SCZ, and underscore the utility of Grin2a-deficient mice as a genetic model of SCZ.

PPM1D mutations are oncogenic drivers of de novo diffuse midline glioma formation.

The role of PPM1D mutations in de novo gliomagenesis has not been systematically explored. Here we analyze whole genome sequences of 170 pediatric high-grade gliomas and find that truncating mutations in PPM1D that increase the stability of its phosphatase are clonal driver events in 11% of Diffuse Midline Gliomas (DMGs) and are enriched in primary pontine tumors. Through the development of DMG mouse models, we show that PPM1D mutations potentiate gliomagenesis and that PPM1D phosphatase activity is required for in vivo oncogenesis. Finally, we apply integrative phosphoproteomic and functional genomics assays and find that oncogenic effects of PPM1D truncation converge on regulators of cell cycle, DNA damage response, and p53 pathways, revealing therapeutic vulnerabilities including MDM2 inhibition.

Profiling Thermus thermophilus Argonaute Guide DNA Sequence Preferences by Functional Screening.

Prokaryotic Argonautes (pAgo) are an increasingly well-studied class of guided endonucleases, and the underlying mechanisms by which pAgo generate nucleic acid guides in vivo remains an important topic of investigation. Recent insights into these mechanisms for the Argonaute protein from Thermus thermophilus has drawn attention to global sequence and structural feature preferences involved in oligonucleotide guide selection. In this work, we approach the study of guide sequence preferences in T. thermophilus Argonaute from a functional perspective. Screening a library of 1,968 guides against randomized single- and double-stranded DNA substrates, endonuclease activity associated with each guide was quantified using high-throughput capillary electrophoresis, and localized sequence preferences were identified which can be used to improve guide design for molecular applications. The most notable preferences include: a strong cleavage enhancement from a first position dT independent of target sequence; a significant decrease in activity with dA at position 12; and an impact of GC dinucleotides at positions 10 and 11. While this method has been useful in characterizing unique preferences of T. thermophilus Argonaute and criteria for creating efficient guides, it could be expanded further to rapidly characterize more recent mesophilic variants reported in the literature and drive their utility toward molecular tools in biology and genome editing applications.