Biology subjects

Stühler, K.

Publications and source records attributed to Stühler, K..

4 recordsLinked to original sources

bioRxiv2025

Canonical autophagy remains inactive in induced pluripotent stem cells and neuronal progenitor cells following DNA damage induced by BPDE or etoposide.

(Macro-)Autophagy is a key cellular stress response mediating the recycling of long-lived or damaged proteins and organelles. In stem cells, autophagy is essential for the decision between quiescence, self-renewal and differentiation. We observed that induced pluripotent stem cells (iPSCs) and thereof derived neural progenitor cells (NPCs) have a functional autophagy machinery, as shown by starvation-induced autophagic flux and ULK1 activation. Using the human iPSC line iPS11 and thereof derived NPCs (niPS11), we investigated whether genotoxic stress induced by benzo[a]pyrene diolepoxide (BPDE) or etoposide can similarly activate autophagy, as previously reported for cancer cell lines. While both BPDE and etoposide induced the DNA damage markers phospho-p53 Ser15 and {gamma}H2AX and slightly altered the expression of DNA repair proteins such as XPC, they did not trigger autophagic flux in either iPSCs or NPCs. After genotoxin treatment, ULK1 activation was only observed in NPCs, but this was not sufficient to trigger a significant downstream autophagic response. Mass spectrometry revealed minimal proteomic changes in iPSCs and moderate changes in NPCs, mainly involving mitotic regulators. These results suggest that genotoxic agents do not strongly affect canonical autophagy in pluripotent stem cells or their neural derivatives despite an otherwise responsive autophagic system.

cell biology↗

bioRxiv2024

Development of a Bipyrimidineamide based α-Helix Mimetic Lead Compound for efficient Targeting of MDM2 in Triple-Negative Breast Cancer

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=155 SRC="FIGDIR/small/582899v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@89ffd0org.highwire.dtl.DTLVardef@1f9cba2org.highwire.dtl.DTLVardef@a821b3org.highwire.dtl.DTLVardef@3c7b6a_HPS_FORMAT_FIGEXP M_FIG C_FIG Triple-negative breast cancer (TNBC) represents the most aggressive form among breast carcinoma subtypes. Due to limited therapy options, identification of novel active pharmacological compounds is an urgent medical need. A promising approach in cancer treatment is the pharmacological inhibition of murine double minutes 2 (MDM2)-p53/p73 interactions inducing apoptosis in tumors. We here describe a novel bipyrimidineamide based -helix mimetic 9 (VWK603) which was designed as a lead candidate to target MDM2. 9 (VWK603) potently induced cell death in the TNBC cell lines MDA-MB-231, MDA-MB-436 and MDA-MB-468 with IC50 values ranging between 3.7 {micro}M and 6.6 {micro}M. The anti-tumor activity was about four more potent higher than determined for the MDM2-specific inhibitor Nutlin-3a. Mechanistic analysis revealed induction of cellular apoptosis as the underlying mode of action of 9 (VWK603) anti-tumor activity. Since toxicity was observed to be reduced in non-cancerous breast cells, these studies make 9 (VWK603) a promising candidate for further preclinical MDM2 inhibitor development.

cancer biology↗

bioRxiv2020

The puzzle of metabolite exchange and identification of putative octotrico peptide repeat expression regulators in the nascent photosynthetic organelles of Paulinella chromatophora

The cercozoan amoeba Paulinella chromatophora contains photosynthetic organelles - termed chromatophores - that evolved from a cyanobacterium, independently from plastids in plants and algae. Despite the more recent origin of the chromatophore, it shows tight integration into the host cell. It imports hundreds of nucleus-encoded proteins, and diverse metabolites are exchanged across the two chromatophore envelope membranes. However, the limited set of chromatophore-encoded transporters appears insufficient for supporting metabolic connectivity or protein import. Furthermore, chromatophore-localized biosynthetic pathways as well as multiprotein complexes include proteins of dual genetic origin, suggesting coordination of gene expression levels between chromatophore and nucleus. These findings imply that similar to the situation in mitochondria and plastids, nuclear factors evolved that control metabolite exchange and gene expression in the chromatophore. Here we show by mass spectrometric analyses of enriched insoluble protein fractions that, unexpectedly, nucleus-encoded transporters are not inserted into the chromatophore inner envelope membrane. Thus, despite the apparent maintenance of its barrier function, canonical metabolite transporters are missing in this membrane. Instead we identified several expanded groups of short chromatophore-targeted orphan proteins. Members of one of these groups are characterized by a single transmembrane helix, and others contain amphipathic helices. We hypothesize that these proteins are involved in modulating membrane permeability. Furthermore, we identified an expanded family of chromatophore-targeted helical repeat proteins. These proteins show similar domain architectures as known organelle-targeted octotrico peptide repeat expression regulators in algae and plants suggesting their convergent evolution as nuclear regulators of gene expression levels in the chromatophore. ImportanceThe endosymbiotic acquisition of mitochondria and plastids >1 billion years ago was central for the evolution of eukaryotic life. However, owing to their ancient origin, these organelles provide only limited insights into the initial stages of organellogenesis. The chromatophore in Paulinella evolved ~100 million years ago and thus, offers the possibility to gain valuable insights into early stages and common rules in organelle evolution. Critical to organellogenesis appears to be the establishment of nuclear control over metabolite exchange and gene expression in the endosymbiont. Here we show that the mechanism generating metabolic connectivity of the chromatophore fundamentally differs from the one for mitochondria and plastids, but likely rather resembles the poorly understood mechanism in various bacterial endosymbionts in plants and insects. Furthermore, we describe a novel class of chromatophore-targeted helical repeat proteins which evolved convergently to plastid-targeted expression regulators and are likely involved in gene expression control in the chromatophore.

evolutionary biology↗

bioRxiv2020

Subcellular localization and mitotic interactome analyses identify SIRT4 as a centrosomally localized and microtubule associated protein

The stress-inducible and senescence-associated tumor suppressor SIRT4, a member of the family of mitochondrial sirtuins (SIRT3, SIRT4, and SIRT5), regulates bioenergetics and metabolism via NAD+-dependent enzymatic activities. Next to the known mitochondrial location, we found that a fraction of endogenous or ectopically expressed SIRT4, but not SIRT3, is located at the mitotic spindle apparatus in the cytosol. Confocal spinning disk microscopy revealed that SIRT4 localizes during the cell cycle dynamically at centrosomes with an intensity peak in G2 and early mitosis. Moreover, SIRT4 binds to microtubules and interacts with structural (,{beta}-tubulin, {gamma}-tubulin, TUBGCP2, TUBGCP3) and regulatory (HDAC6) microtubule components as detected by co-immunoprecipitation and mass spectrometric analyses of the mitotic SIRT4 interactome. Overexpression of SIRT4 resulted in a pronounced decrease of acetylated -tubulin (K40) associated with altered microtubule dynamics in mitotic cells. SIRT4 or the N-terminally truncated variant SIRT4({Delta}N28), which is unable to translocate into mitochondria, delayed mitotic progression and reduced cell proliferation. This study extends the functional roles of SIRT4 beyond mitochondrial metabolism, and suggests that SIRT4 acts as a novel centrosomal / microtubule-associated protein in the regulation of cell cycle progression. Thus, stress-induced SIRT4 may exert its role as tumor suppressor through mitochondrial as well as extramitochondrial functions, the latter associated with its localization at the mitotic spindle apparatus.

cell biology↗