Biology subjects

Linder, S.

Publications and source records attributed to Linder, S..

9 recordsLinked to original sources

bioRxiv2026

Drebrin forms a cortical hub that connects actin, microtubules, and clathrin for endocytic transport of β2 integrin

Using primary human macrophages, we analyze the molecular machinery that enables direct contact of microtubule +tips with podosomes, and its connection to clathrin-based endocytosis. We show that the podosome cap protein drebrin, together with its +tip-localized binding partner EB3, regulates podosome-microtubule contact. Importantly, drebrin depletion leads to reduced endocytosis, particularly of {beta}2 integrin at podosomes. This is based on drebri[n]s interaction with {beta}2 integrin, and its direct binding of clathrin heavy chain via a canonical clathrin box and a novel box, which we identify also in other mammalian proteins. Our data enable a model of the molecular machinery that regulates direct contact between podosomes and microtubule +tips. Moreover, we provide evidence for spatial selectivity in endocytosis of {beta} integrins, while also revealing a more general principle of endocytic trafficking regulation, through drebrin working as a multivalent, actin-based hub that links the clathrin system with microtubule-based transport.

cell biology↗

bioRxiv2025

Phagocytic podosomes enable efficient uptake of Candida auris by primary human macrophages

The yeast Candida auris is an emerging pathogen of steadily increasing importance. Understanding the molecular mechanisms of C. auris uptake and intracellular processing by immune cells such as macrophages is thus critical for counteracting the spreading of respective infections. Here, we show that phagocytosis of C. auris cells by primary human macrophages involves the formation of dot-like F-actin-rich structures at C. auris-containing phagosomes that we characterize as phagocytic podosomes. We analyse the composition, architecture, and dynamics of these structures, showing that they constitute a specific and highly dynamic adaptation of the phagocytic actin network of macrophages. We also show that disruption of phagocytic podosomes is associated with reduced internalization of C. auris cells and delayed maturation of respective phagosomes. Our data provide novel insights into the uptake mechanism and cytoskeletal rearrangements upon internalization of Candida by immune cells, while also challenging the dogma of a generally uniform and continuous actin network within phagocytic cups. At the same time, we identify C. auris as the first pathophysiologically relevant target whose internalization involves the formation of phagocytic podosomes.

cell biology↗

bioRxiv2025

Myo1e/f at the podosome base regulate podosome dynamics and promote macrophage migration

Cells of the monocyte lineage form specialized membrane-associated, actin-rich structures, called podosomes. Podosomes play important roles in cell adhesion and migration as well as the proteolytic degradation of the extracellular matrix. While podosomes are always closely associated with the plasma membrane, the structural components linking the podosome core, composed of branched actin, to the membrane are not fully understood. In this study we show that class I myosins, Myo1e and Myo1f, localize to a specific region of podosomes, underneath the podosome core and near the ventral plasma membrane, and that this localization is mainly mediated by the Myo1e/f TH2 domains. Respective knockdowns or knockouts of Myo1e/f lead to increased podosome size, altered turnover and lateral mobility, which is likely due to Myo1e/f regulating the attachment of core actin filaments to the plasma membrane. In addition, Myo1e/f double knockout macrophages were characterized by a reduction in 3D and 2D migration, even though these cells exhibited increased ability to degrade the extracellular matrix. Along with the other membrane-associated podosome components, such as the transmembrane proteins MT-MMP and CD44, and the GPI-anchored DNase X, Myo1e and Myo1f mark the membrane-proximal region of podosomes. We propose to label this region as the podosome "base", an additional substructure joining the current trifecta of the podosome cap, core, and ring.

cell biology↗

bioRxiv2025

Discovery of a new evolutionarily conserved short linear F-actin binding motif

Regulation of the actin cytoskeleton by actin binding proteins (ABPs) is essential for cellular homeostasis, and the mode of actin binding determines the activity of ABPs. Here, we discovered a novel "Short linear F-actin binding motif (SFM)" on the basis of the cryo-EM structure of the ITPKA-F-actin complex. We developed the computational pipeline SLiMFold, which identified 103 human SFM containing-proteins exhibiting diverse cellular functions. The SFM probably developed ex nihilo and remained conserved in eukaryotes, with a binding affinity to F-actin ranging from 13 to 89 {micro}M. Furthermore, we uncovered the essential amino acids of this SFM for F-actin binding and affinity modulation. Together, the SFM seems to serve as a low affinity anchor to target proteins to F-actin, in order to connect the regulation of actin dynamics with broad cellular functions. These findings will shed new light on the role of a wide variety of proteins.

biochemistry↗

bioRxiv2025

TRIM33 loss reduces Androgen Receptor transcriptional output and H2BK120 ubiquitination

The Androgen Receptor (AR) is a ligand-dependent transcription factor that drives prostate cancer development and progression. Although, a detailed effect on AR biology has been described for a number of interacting proteins, many AR coregulators remain to be characterized in relation to their distinct impact on AR function. Here, we describe TRIM33 as a conserved AR-interactor across multiple prostate cancer cell lines. We observed that TRIM33 and AR share overall chromatin interaction profiles, in which TRIM33 is involved in downstream responsive transcriptomic output. In contrast to prior reports, we show that TRIM33 does not impact AR protein stability, but instead propose a model in which TRIM33 facilitates maximal AR activity by interfering with H2BK120 ubiquitination levels.

molecular biology↗

bioRxiv2021

Genetic determinants of chromatin reveal prostate cancer risk mediated by context-dependent gene regulation

Methods that link genetic variation to steady-state gene expression levels, such as expression quantitative trait loci (eQTLs), are widely used to functionally annotate trait-associated variants, but they are limited in identifying context-dependent effects on transcription. To address this challenge, we developed the cistrome-wide association study (CWAS), a framework for nominating variants that impact traits through their effects on chromatin state. CWAS associates the genetic determinants of cistromes (e.g., the genome-wide profiles of transcription factor binding sites or histone modifications) with traits using summary statistics from genome-wide association studies (GWAS). We performed CWASs of prostate cancer and androgen-related traits, using a reference panel of 307 prostate cistromes from 165 individuals. CWAS nominated susceptibility regulatory elements or androgen receptor (AR) binding sites at 52 out of 98 known prostate cancer GWAS loci and implicated an additional 17 novel loci. We functionally validated a subset of our results using CRISPRi and in vitro reporter assays. At 28 of the 52 risk loci, CWAS identified regulatory mechanisms that are not observable via eQTLs, implicating genes with complex or context-specific regulation that are overlooked by current approaches that relying on steady-state transcript measurements. CWAS genes include transcription factors that govern prostate development such as NKX3-1, HOXB13, GATA2, and KLF5. Moreover, CWAS boosts discovery power in modestly sized GWAS, identifying novel genetic associations mediated through AR binding for androgen-related phenotypes, including resistance to prostate cancer therapy. CWAS is a powerful and biologically interpretable paradigm for studying variants that influence traits by affecting context-dependent transcriptional regulation.

genetics↗

bioRxiv2021

Targeting Mitochondrial Metabolism in Clear Cell Carcinoma of the Ovaries

Ovarian clear cell carcinoma (OCCC) is a rare but chemorefractory tumor. About 50% of all OCCC patients have inactivating mutations of ARID1A a member of the SWI/SNF chromatin remodeling complex. Members of the SWI/SNF remodeling have emerged as regulators of the energetic metabolism of mammalian cells, however the role of ARID1A as a modulator of the mitochondrial metabolism in OCCCs is yet to be defined. Here we show that ARID1A-loss results in increased mitochondrial metabolism and renders ARID1A-mutated cells increasingly and selectively dependent on it. The increase in mitochondrial activity following ARID1A loss is associated to increase of C-myc and to increased mitochondrial number and reduction of their size consistent with a higher mitochondrial cristae/outer membrane ratio. Significantly, preclinical testing of the complex I mitochondrial inhibitor IACS-010759 extends overall survival in a preclinical model of ARID1A-mutated OCCC. These findings provide the for targeting mitochondrial activity in ARID1A-mutanted OCCCs.

cancer biology↗

bioRxiv2020

Cargo-specific recruitment in clathrin and dynamin-independent endocytosis

Spatially controlled, cargo-specific endocytosis is essential for development, tissue homeostasis, and cancer invasion and is often hijacked by viral infections 1. Unlike clathrin-mediated endocytosis, which exploits cargo-specific adaptors for selective protein internalization, the clathrin and dynamin-independent endocytic pathway (CLIC-GEEC, CG-pathway) has until now been considered a bulk internalization route for the fluid phase, glycosylated membrane proteins and lipids 2,3. Although the core molecular players of CG endocytosis have been recently defined, no cargo-specific adaptors are known and evidence of selective protein uptake into the pathway is lacking 3. Here, we identify the first cargo-specific adaptor for CG-endocytosis and demonstrate its clinical relevance in breast cancer progression. By combining unbiased molecular characterization and super-resolution imaging, we identified the actin-binding protein swiprosin-1 (EFHD2) as a cargo-specific adaptor regulating integrin internalization via the CG-pathway. Swiprosin-1 couples active Rab21-associated integrins with key components of the CG-endocytic machinery, IRSp53 and actin. Swiprosin-1 is critical for integrin endocytosis, but not for other CG-cargo and supports integrin-dependent cancer cell migration and invasion, with clinically relevant implications for breast cancer. Our results demonstrate a previously unknown cargo selectivity for the CG-pathway and opens the possibility to discover more adaptors regulating it.

cell biology↗

bioRxiv2020

Functional mapping of androgen receptor enhancer activity

Androgen receptor (AR) is critical to the initiation, growth and progression of almost all prostate cancers. Once activated, the AR binds to cis-regulatory enhancer elements on DNA that drive gene expression. Yet, there are 10-100x more binding sites than differentially expressed genes. It still remains unclear how individual sites contribute to AR-mediated transcription. While descriptive functional genomic approaches broadly correlate with enhancer activity, they do not provide the locus-specific resolution needed to delineate the underlying regulatory logic of AR-mediated transcription. Therefore, we functionally tested all commonly occuring clinical AR binding sites with Self-Transcribing Active Regulatory Regions sequencing (STARRseq) to generate the first map of intrinsic AR enhancer activity. This approach is not significantly affected by endogenous chromatin modifications and measures the potential enhancer activity at each cis-regulatory element. Interestingly we found that only 7% of AR binding sites displayed increased enhancer activity upon hormonal stimulation. Instead, the vast majority of AR binding sites were either inactive (81%) or constitutively active enhancers (11%). These annotations strongly correlated with enhancer-associated features in both cell line and clinical prostate cancer. With these validated annotations we next investigated the effect of each enhancer class on transcription and found that AR-driven inducible enhancers frequently interacted with promoters, forming central chromosomal loops critical for gene transcription. We demonstrated that these inducible enhancers act as regulatory hubs that increase contacts with both other AR binding sites and gene promoters. This functional map was used to identify a somatic mutation that significantly reduces the expression of a commonly mutated AR-regulated tumour suppressor. Together, our data reveal a complex interplay between different AR binding sites that work in a highly coordinated manner to drive gene transcription.

cancer biology↗