Stem cells have the unique ability to undergo asymmetric division which produces two daughter cells that are genetically identical, but commit to different cell fates. The loss of this balanced asymmetric outcome can lead to many diseases, including cancer and tissue dystrophy. Understanding this tightly regulated process is crucial in developing methods to treat these abnormalities. Here, we report that during a Drosophila female germline stem cell asymmetric division, the two daughter cells differentially inherit histones at key genes related to either maintaining the stem cell state or promoting differentiation, but not at constitutively active or silenced genes. We combine histone labeling with DNA Oligopaints to distinguish old versus new histones and visualize their inheritance patterns at a single-gene resolution in asymmetrically dividing cells in vivo. This strategy can be applied to other biological systems involving cell fate change during development or tissue homeostasis in multicellular organisms.

Stem cells have the unique ability to undergo asymmetric division which produces two daughter cells that are genetically identical, but commit to different cell fates. The loss of this balanced asymmetric outcome can lead to many diseases, including cancer and tissue dystrophy. Understanding this tightly regulated process is crucial in developing methods to treat these abnormalities. Here, we report that produced from a Drosophila female germline stem cell asymmetric division, the two daughter cells differentially inherit histones at key genes related to either maintaining the stem cell state or promoting differentiation, but not at constitutively active or silenced genes. We combined histone labeling with DNA Oligopaints to distinguish old versus new histone distribution and visualize their inheritance patterns at single-gene resolution in asymmetrically dividing cells in vivo. This strategy can be widely applied to other biological contexts involving cell fate establishment during development or tissue homeostasis in multicellular organisms.

BACKGROUND Older age is a poor prognostic factor for glioblastoma (GBM) patients. We tested whether the intrinsic molecular landscape of the tumor may contribute to this poor prognosis. METHODS In accordance with the 2021 WHO classification scheme, we included only isocitrate dehydrogenase (IDH) wild type GBM. Based on published literature, we defined older as age > 65. RNA expression, gene amplification, tumor mutational burden (TMB) and mutational profiles were analyzed in three unique datasets: Tempus (n = 1,410), Caris (n = 1,432), and TCGA (n = 557). Comparison were made between < 65 and ³ 65 year olds using Pearson’s Chi-squared tests, Fisher’s exact tests, or Wilcoxon rank-sum where appropriate. RESULTS From our evaluable gene sets, TERT promoter mutations were more prevalent in patients ³ 65 years old (Caris 82.64 vs 77.27%, p = 0.016; Tempus 58.0 vs 49.0%, p = 0.002). There were no significant differences in PDCD1, CD274, CD3E, TNFRSF18, CD40, CD8A, TNFRSF4, CTLA4, HAVCR2, TNFSF9, CD274, or CDKN2A; PDL-1 (by IHC); dMMR/MSI-H, TMB; CDK6 amplification, EGFR amplification, EGFR, EGFRvIII, EGFR fusions, MET fusions, PTEN, TP53, or NF-1. MGMT promoter methylation (Caris data) was more common in the older group (49.73 v 34.14%, p < 0.001). TGCA data demonstrated that gene expression, TMB, and methylation did not change significantly with age. Additionally, PCOLCE2 and SLC10A4 were differentially methylated, and missense mutations, of any type, were more common in the older group (p=0.006). CONCLUSION Despite worse survival outcomes for older patients with IDHwt GBM as compared to younger counterparts, the molecular landscape is similar at the genomic, transcriptomic and epigenomic levels. The key exception is TERT promoter mutations that are more common in older GBM patients. Poorer survival is therefore not likely to be attributable solely to intratumoral factors.

DNA elements act across long genomic distances to regulate gene expression in processes including enhancer-promoter interactions and imprinting. During the gene-regulatory phenomenon of transvection in Drosophila, DNA elements on one allele of a gene act between chromosomes to increase or decrease expression of another allele of the gene. Despite the discovery of transvection over 60 years ago, little is known about its biological role. Furthermore, how different cis regulatory DNA elements contribute to the activation or repression of transvection at distinct times during development is unclear. Here, we studied the stochastic expression of spineless (ss) in developing photoreceptors in the fly eye to understand gene activation and repression between chromosomes. We identified a biological role for transvection in regulating expression of naturally occurring ss alleles. We characterized CRISPR-engineered deletions of sequences across the ss locus and identified DNA elements required for activating and repressing transvection. We found that different enhancers participated in transvection at different times during development to promote gene expression and specify cell fates. Bringing a silencer element on a heterologous chromosome into proximity with the ss locus reconstituted the gene, leading to repression. Our studies show that transvection regulates gene expression via distinct DNA elements at specific timepoints in development, with implications for genome organization and architecture.