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Wippich, F.

Publications and source records attributed to Wippich, F..

4 recordsLinked to original sources

Granule microenvironment regulates the dual functions of FMR1

Fragile X Messenger Ribonucleoprotein 1 (FMR1) is an evolutionarily conserved RNA binding protein with important functions in cognition and female reproduction, and its disruption is associated with neurodevelopmental and reproductive disorders including the Fragile X syndrome. FMR1 is best known for its role as a translation repressor. However, several recent studies also suggest a role of FMR1 as a translation enhancer raising fundamental questions about the molecular regulation of these opposing functions. In this study, we identify FMR1 as part of the oskar mRNA-protein complex in the Drosophila oocyte and study the role of FMR1 as a translational enhancer of oskar. We provide the molecular mechanism for the dual functions of FMR1 and show that the two major RNA-binding domains of FMR1, the KH domains and the RGG box, play distinct separable roles in regulating translation. The KH domains enhance translation of mRNAs while the RGG box containing C-terminal domain (CTD) is required to repress translation. We further show that the condensation propensity of FMR1 containing granules regulates the two antagonistic functions, such that phase separation by FMR1-CTD creates the molecular microenvironment necessary for the repressive activity, whereas reduction in phase separation is associated with increased translation. Our findings highlight the importance of biomolecular condensates not just as a means of molecular compartmentalization but as a fundamental regulatory principle that dictates the functional output of modular protein domains.

molecular biology↗

iss-nf: A Nextflow-based end-to-end in situ sequencing decoding workflow

In situ sequencing (ISS) offers a powerful approach for spatially resolved gene expression profiling within tissue samples, but the complexity of analyzing the resulting data has limited its broader use. Here, we present iss-nf, a Nextflow-based, end-to-end workflow designed to streamline the decoding of large ISS datasets. The workflow automates critical steps, including image registration, fluorescent spot detection, transcript decoding, and quality control (QC), offering a scalable, reproducible, and user-friendly solution for ISS data analysis. We successfully applied iss-nf on multiple large datasets, including publicly available mouse brain and breast cancer tissue datasets, as well as an in-house non-small cell lung cancer (NSCLC) ISS dataset. The workflow is designed to be accessible for both experienced researchers, as well as newcomers to spatial transcriptomics, providing a robust tool for analyzing large-scale ISS data. Our results suggest that iss-nf is a valuable contribution to the growing field of spatial transcriptomics, enabling precise, modular, reproducible, and, by means of automated tiling and parallelization, scalable analysis of tissue-specific gene expression.

bioinformatics↗

Nutritional stress-induced regulation of microtubule organization and mRNP transport by HDAC1 controlled α-tubulin acetylation

In response to nutritional stress, microtubules in cells of the Drosophila female germline are depleted from the cytoplasm and accumulate cortically. This triggers aggregation of mRNPs into large reticulated sponge bodies and oogenesis arrest. Here, we show that hyperacetylation of -tubulin at lysine 40 (K40) alters microtubule dynamics and sponge body formation. We found that depletion of histone deacetylase 1 (HDAC1) by RNAi phenocopies the nutritional stress response, causing hyperacetylation of -tubulin, as well as accumulation of maternally deposited mRNPs in reticulated sponge bodies. Using in vitro and in vivo studies, we identify HDAC1 as a direct regulator of -tubulin K40 acetylation status. In well-fed flies, HDAC1 maintains low levels of -tubulin acetylation, enabling the microtubule dynamics required for mRNP transport. Using quantitative phosphoproteomics we identify nutritional stress-induced changes in protein phosphorylation that act upstream of -tubulin acetylation, including phosphorylation of HDAC1 at S391, which reduces its ability to deacetylate -tubulin. These results reveal that Drosophila HDAC1 senses and relays the nutritional status, which regulates germline development through modulation of cytoskeleton dynamics.

developmental biology↗

Enzymatic production of single molecule FISH and RNA capture probes

Arrays of singly-labelled short oligonucleotides that hybridize to a specific target revolutionized RNA biology, enabling quantitative, single molecule microscopy analysis and high efficiency RNA/RNP capture. Here, we describe a simple and efficient method that allows flexible functionalization of inexpensive DNA oligonucleotides by different fluorescent dyes or biotin using terminal deoxynucleotidyl transferase and custom-made functional group conjugated dideoxy-UTP. We show that 1) all steps of the oligonucleotide labelling - including conjugation, enzymatic synthesis and product purification - can be performed in a standard biology laboratory, 2) the process yields >90 %, often >95 % labeled product with minimal carry-over of impurities and 3) the oligonucleotides can be labeled with different dyes or biotin, allowing single molecule FISH or RNA affinity purification to be performed.\n\nGraphical abstract\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=66 SRC=\"FIGDIR/small/107599_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (20K):\norg.highwire.dtl.DTLVardef@a26162org.highwire.dtl.DTLVardef@a780d9org.highwire.dtl.DTLVardef@fc3a62org.highwire.dtl.DTLVardef@15d183c_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗