Biology subjects

Krabbenhoft, S. D.

Publications and source records attributed to Krabbenhoft, S. D..

3 recordsLinked to original sources

bioRxiv2025

Investigating oncoprotein-mediated chromatin dysregulation in Drosophila melanogaster uncovers novel modifiers of the developmental impact of H3 K27M and EZHIP

Polycomb Repressive Complex 2 (PRC2) maintains epigenetic repression through the catalysis of H3K27 trimethylation (H3K27me3), which restricts gene expression and preserves developmental gene-regulatory networks. The integrity of PRC2-mediated gene silencing depends critically on the ability of PRC2 to establish and propagate H3K27me3 beyond initial recruitment sites. The oncoproteins EZHIP and histone H3 K27M specifically inhibit this propagation by blocking the allosterically activated state of PRC2, leading to global disruption of H3K27me3 patterns and developmental abnormalities. To uncover chromatin-related pathways intersecting with PRC2 repression, we developed a Drosophila melanogaster model with tissue-specific expression of EZHIP and H3 K27M. A targeted RNAi screen of conserved chromatin regulators identified genetic modifiers that when knocked down either enhanced or suppressed developmental phenotypes driven by these PRC2 inhibitors. Strong suppressors, including the Trithorax-group proteins Ash1 and Trx, the PR-DUB complex member Asx, and the nucleoporin Nup153, restored normal development despite persistent depletion of global H3K27me3. Gene expression analyses revealed that suppression reflected reduced expression of genes aberrantly activated following PRC2 inhibition. Together, these findings highlight conserved chromatin-regulatory pathways that intersect with Polycomb to maintain transcriptional balance and support developmental homeostasis.

developmental biology↗

bioRxiv2022

Premature translation of the zygotic genome activator Zelda is not sufficient to precociously activate gene expression

Following fertilization, the unified germ cells rapidly transition to a totipotent embryo. Maternally deposited mRNAs encode the proteins necessary for this reprogramming as the zygotic genome remains transcriptionally quiescent during the initial stages of development. The transcription factors required to activate the zygotic genome are among these maternally deposited mRNAs and are robustly translated following fertilization. In Drosophila, the mRNA encoding Zelda, the major activator of the zygotic genome, is not translated until one hour after fertilization. Here we demonstrate that zelda translation is repressed following fertilization by the TRIM-NHL protein Brain tumor (BRAT). BRAT also regulates Zelda levels in the larval neuroblast lineage. In the embryo, BRAT-mediated translational repression is regulated by the Pan Gu (PNG) kinase, which is triggered by egg activation. The PNG kinase phosphorylates translational regulators, suggesting that PNG kinase activity alleviates translational repression of zelda by BRAT and coupling translation of zelda with that of other regulators of early embryonic development. Using the premature translation of zelda in embryos lacking BRAT activity, we showed that early translation of a zygotic genome activator is not sufficient to drive precocious gene expression. Instead, Zelda-target genes showed increased expression at the time they are normally activated. We propose that transition through early development requires the integration of multiple processes, including the slowing of the nuclear division cycle and activation of the zygotic genome. These processes are coordinately controlled by PNG kinase-mediated regulation of translation.

developmental biology↗

bioRxiv2020

H3 K27M and EZHIP impede H3K27-methylation spreading by inhibiting allosterically stimulated PRC2

Diffuse midline gliomas and posterior fossa type-A ependymomas contain the highly recurrent histone H3 K27M mutation and the H3 K27M-mimic EZHIP, respectively. In vitro, H3 K27M and EZHIP are competitive inhibitors of Polycomb Repressive Complex 2 (PRC2) lysine methyltransferase activity. In vivo, these proteins reduce overall H3K27me3 levels, however residual peaks of H3K27me3 remain at CpG islands through an unknown mechanism. Here, we report that EZHIP and H3 K27M preferentially interact with an allosterically activated form of PRC2 in vivo. The formation of H3 K27M- and EZHIP-PRC2 complexes occurs at CpG islands containing H3K27me3 and impedes PRC2 and H3K27me3 spreading. While EZHIP is not found outside of placental mammals, we find that expression of human EZHIP reduces H3K27me3 in Drosophila melanogaster through a conserved molecular mechanism. Our results highlight the mechanistic similarities between EZHIP and H3 K27M in vivo and provide mechanistic insight for the retention of residual H3K27me3 in tumors driven by these oncogenes.

cancer biology↗