Altered neuroepithelial morphogenesis and migration defects in iPSC-derived cerebral organoids and 2D neural stem cells in familial bipolar disorder.

Bipolar disorder (BD) is a severe mental illness that can result from neurodevelopmental aberrations, particularly in familial BD, which may include causative genetic variants. In the present study, we derived cortical organoids from BD patients and healthy (control) individuals from a clinically dense family in the Indian population. Our data reveal that the patient organoids show neurodevelopmental anomalies, including organisational, proliferation and migration defects. The BD organoids show a reduction in both the number of neuroepithelial buds/cortical rosettes and the ventricular zone size. Additionally, patient organoids show a lower number of SOX2-positive and EdU-positive cycling progenitors, suggesting a progenitor proliferation defect. Further, the patient neurons show abnormal positioning in the ventricular/intermediate zone of the neuroepithelial bud. Transcriptomic analysis of control and patient organoids supports our cellular topology data and reveals dysregulation of genes crucial for progenitor proliferation and neuronal migration. Lastly, time-lapse imaging of neural stem cells in 2D in vitro cultures reveals abnormal cellular migration in BD samples. Overall, our study pinpoints a cellular and molecular deficit in BD patient-derived organoids and neural stem cell cultures.

LSD1 Regulates Neurogenesis in Human Neural Stem Cells Through the Repression of Human-Enriched Extracellular Matrix and Cell Adhesion Genes.

Neurogenesis begins with neural stem cells undergoing symmetric proliferative divisions to expand and then switching to asymmetric differentiative divisions to generate neurons in the developing brain. Chromatin regulation plays a critical role in this switch. Histone lysine-specific demethylase LSD1 demethylates H3K4me1/2 and H3K9me1/2 but the mechanisms of its global regulatory functions in human neuronal development remain unclear. We performed genome-wide ChIP-seq of LSD1 occupancy, RNA-seq, and Histone ChIP-seq upon LSD1 inhibition to identify its repressive role in human neural stem cells. Novel downstream effectors of LSD1 were identified, including the Notch signaling pathway genes and human-neural progenitor-enriched extracellular matrix (ECM) pathway/cell adhesion genes, which were upregulated upon LSD1 inhibition. LSD1 inhibition led to decreased neurogenesis, and overexpression of downstream effectors mimicked this effect. Histone ChIP-seq analysis revealed that active and enhancer markers H3K4me2, H3K4me1, and H3K9me1 were upregulated upon LSD1 inhibition, while the repressive H3K9me2 mark remained mostly unchanged. Our work identifies the human-neural progenitor-enriched ECM pathway/cell adhesion genes and Notch signaling pathway genes as novel downstream effectors of LSD1, regulating neuronal differentiation in human neural stem cells.

Disrupting butterfly caterpillar microbiomes does not impact their survival and development.

Associations with gut microbes are believed to play crucial roles in the physiology, immune function, development and behaviour of insects. However, microbiome sequencing has recently suggested that butterflies are an anomaly, because their microbiomes do not show strong host- and developmental stage-specific associations. We experimentally manipulated butterfly larval gut microbiota and found that disrupting gut microbes had little influence on larval survival and development. Larvae of the butterflies Danaus chrysippus and Ariadne merione that fed on chemically sterilized or antibiotic-treated host plant leaves had significantly reduced bacterial loads, and their gut bacterial communities were disrupted substantially. However, neither host species treated this way suffered a significant fitness cost: across multiple experimental blocks, treated and control larvae had similar survival, growth and development. Furthermore, re-introducing microbes from the excreta of control larvae did not improve larval growth and survival. Thus, these butterfly larvae did not appear to rely on specialized gut bacteria for digestion, detoxification, biomass accumulation and metamorphosis. Our experiments thus show that dependence on gut bacteria for growth and survival is not a universal phenomenon across insects. Our findings also caution that strategies which target gut microbiomes may not always succeed in controlling Lepidopteran pests.

Dietary and developmental shifts in butterfly-associated bacterial communities.

Bacterial communities associated with insects can substantially influence host ecology, evolution and behaviour. Host diet is a key factor that shapes bacterial communities, but the impact of dietary transitions across insect development is poorly understood. We analysed bacterial communities of 12 butterfly species across different developmental stages, using amplicon sequencing of the 16S rRNA gene. Butterfly larvae typically consume leaves of a single host plant, whereas adults are more generalist nectar feeders. Thus, we expected bacterial communities to vary substantially across butterfly development. Surprisingly, only few species showed significant dietary and developmental transitions in bacterial communities, suggesting weak impacts of dietary transitions across butterfly development. On the other hand, bacterial communities were strongly influenced by butterfly species and family identity, potentially due to dietary and physiological variation across the host phylogeny. Larvae of most butterfly species largely mirrored bacterial community composition of their diets, suggesting passive acquisition rather than active selection. Overall, our results suggest that although butterflies harbour distinct microbiomes across taxonomic groups and dietary guilds, the dramatic dietary shifts that occur during development do not impose strong selection to maintain distinct bacterial communities across all butterfly hosts.

Large-Effect Beneficial Synonymous Mutations Mediate Rapid and Parallel Adaptation in a Bacterium.

Contrary to previous understanding, recent evidence indicates that synonymous codon changes may sometimes face strong selection. However, it remains difficult to generalize the nature, strength, and mechanism(s) of such selection. Previously, we showed that synonymous variants of a key enzyme-coding gene (fae) of Methylobacterium extorquens AM1 decreased enzyme production and reduced fitness dramatically. We now show that during laboratory evolution, these variants rapidly regained fitness via parallel yet variant-specific, highly beneficial point mutations in the N-terminal region of fae These mutations (including four synonymous mutations) had weak but consistently positive impacts on transcript levels, enzyme production, or enzyme activity. However, none of the proposed mechanisms (including internal ribosome pause sites or mRNA structure) predicted the fitness impact of evolved or additional, engineered point mutations. This study shows that synonymous mutations can be fixed through strong positive selection, but the mechanism for their benefit varies depending on the local sequence context.

Cell surfactomes of two endometrial epithelial cell lines that differ in their adhesiveness to embryonic cells.

Adhesiveness of the endometrial epithelium to an embryo plays a critical role in the initiation of pregnancy. Loss or gain of adhesiveness also dictates the potential of endometrial epithelial cells to metastasize, an event that can result from certain genetic insults. A proteomics-based exploration of the "adhesiveness" these epithelial cells was employed that could identify targets that could disrupt embryo-endometrium interactions and/or metastasis of endometrial cancer cells. The present study defined the surfactomes of two human endometrial epithelial cell lines known for their differential adhesiveness to embryonic cells. Comparative, two-dimensional electrophoretic analysis of the surfactomes of RL95-2 (exhibiting higher adhesiveness to the embryonic cell line JAr) and HEC-1A (exhibiting reduced adhesiveness to JAr cells) revealed 55 differentially enriched proteins. Of these, 10 proteins were identified by MALDI-TOF/TOF or LC-MS/MS. TUBB2C, ADAMTS3, and elongation factor beta were more abundant on the HEC-1A cell surface whereas HSP27, HSPA9, GP96, CRT, Tapasin-ERP57, PDI, and ß-actin were more abundant on the RL95-2 cell surface. Nano LC-MS/MS was also employed to generate a more comprehensive surfactomes of RL95-2 and HEC-1A. The study also demonstrated a pro-adhesive role of CRT and HSPA9 and an anti-adhesive role of TUBB2C populations found on the cell surface. In brief, this study identifies the cell-surface protein complements of two human endometrial epithelial cell lines, and reveals the role of three proteins in heterotypic cell adhesion.