A new dawn for the study of cell type evolution.

Animal evolution is influenced by the emergence of new cell types, yet our understanding of this process remains elusive. This prompts the need for a broader exploration across diverse research organisms, facilitated by recent breakthroughs, such as gene editing tools and single-cell genomics. Essential to our understanding of cell type evolution is the accurate identification of homologous cells. We delve into the significance of considering developmental ontogeny and potential pitfalls when drawing conclusions about cell type homology. Additionally, we highlight recent discoveries in the study of cell type evolution through the application of single-cell transcriptomics and pinpoint areas ripe for further exploration.

Proteostasis governs differential temperature sensitivity across embryonic cell types.

Embryonic development is remarkably robust, but temperature stress can degrade its ability to generate animals with invariant anatomy. Phenotypes associated with environmental stress suggest that some cell types are more sensitive to stress than others, but the basis of this sensitivity is unknown. Here, we characterize hundreds of individual zebrafish embryos under temperature stress using whole-animal single-cell RNA sequencing (RNA-seq) to identify cell types and molecular programs driving phenotypic variability. We find that temperature perturbs the normal proportions and gene expression programs of numerous cell types and also introduces asynchrony in developmental timing. The notochord is particularly sensitive to temperature, which we map to a specialized cell type: sheath cells. These cells accumulate misfolded protein at elevated temperature, leading to a cascading structural failure of the notochord and anatomic defects. Our study demonstrates that whole-animal single-cell RNA-seq can identify mechanisms for developmental robustness and pinpoint cell types that constitute key failure points.

MS4A4A modifies the risk of Alzheimer disease by regulating lipid metabolism and immune response in a unique microglia state.

Genome-wide association studies (GWAS) have identified many modifiers of Alzheimer disease (AD) risk enriched in microglia. Two of these modifiers are common variants in the MS4A locus (rs1582763: protective and rs6591561: risk) and serve as major regulators of CSF sTREM2 levels. To understand their functional impact on AD, we used single nucleus transcriptomics to profile brains from carriers of these variants. We discovered a "chemokine" microglial subpopulation that is altered in MS4A variant carriers and for which MS4A4A is the major regulator. The protective variant increases MS4A4A expression and shifts the chemokine microglia subpopulation to an interferon state, while the risk variant suppresses MS4A4A expression and reduces this subpopulation of microglia. Our findings provide a mechanistic explanation for the AD variants in the MS4A locus. Further, they pave the way for future mechanistic studies of AD variants and potential therapeutic strategies for enhancing microglia resilience in AD pathogenesis.

A CRISPRi/a platform in human iPSC-derived microglia uncovers regulators of disease states.

Microglia are emerging as key drivers of neurological diseases. However, we lack a systematic understanding of the underlying mechanisms. Here, we present a screening platform to systematically elucidate functional consequences of genetic perturbations in human induced pluripotent stem cell-derived microglia. We developed an efficient 8-day protocol for the generation of microglia-like cells based on the inducible expression of six transcription factors. We established inducible CRISPR interference and activation in this system and conducted three screens targeting the 'druggable genome'. These screens uncovered genes controlling microglia survival, activation and phagocytosis, including neurodegeneration-associated genes. A screen with single-cell RNA sequencing as the readout revealed that these microglia adopt a spectrum of states mirroring those observed in human brains and identified regulators of these states. A disease-associated state characterized by osteopontin (SPP1) expression was selectively depleted by colony-stimulating factor-1 (CSF1R) inhibition. Thus, our platform can systematically uncover regulators of microglial states, enabling their functional characterization and therapeutic targeting.

Functional regulatory variants implicate distinct transcriptional networks in dementia.

Predicting the function of noncoding variation is a major challenge in modern genetics. In this study, we used massively parallel reporter assays to screen 5706 variants identified from genome-wide association studies for both Alzheimer's disease (AD) and progressive supranuclear palsy (PSP), identifying 320 functional regulatory variants (frVars) across 27 loci, including the complex 17q21.31 region. We identified and validated multiple risk loci using CRISPR interference or excision, including complement 4 (C4A) and APOC1 in AD and PLEKHM1 and KANSL1 in PSP. Functional variants disrupt transcription factor binding sites converging on enhancers with cell type-specific activity in PSP and AD, implicating a neuronal SP1-driven regulatory network in PSP pathogenesis. These analyses suggest that noncoding genetic risk is driven by common genetic variants through their aggregate activity on specific transcriptional programs.