Biology subjects

Rothlin, C. V.

Publications and source records attributed to Rothlin, C. V..

8 recordsLinked to original sources

bioRxiv2024

Functional roles of neural aPKCs in mouse brain development and survival

Conserved protein complexes establish and maintain cell polarity. In turn, cell polarity is indispensable for fundamental developmental processes such as asymmetric division of stem cells and establishment of subcellular membrane polarization during cell differentiation. There are three well characterized polarity complexes. Atypical Protein Kinase C (aPKC) is a conserved constituent of the PAR complex that phosphorylates not only substrates within this complex, but also in the other polarity complexes. Outside of the polarity complex, aPKC regulates a myriad of cellular processes such as migration, metabolism, and survival. In mammals, two paralogs, Prkci and Prkcz, form the aPKC subfamily. Here, we characterized the expression of the Prkci and Prkcz paralogs, including a variant transcript of Prkcz, in the mouse brain and specific cells of the neural lineage. We generated a series of mice with individual and collective ablation of the two aPKC paralogs in neural stem cells, new-born neurons, astrocytes and NG2+ cells, as well as a mouse expressing kinase-inactive PRKCI in neural progenitors. We examine the effects of loss of aPKC paralogs or its kinase activity on gross brain development and organismal viability. Our results identify a critical window in neural progenitor differentiation wherein aPKC function is indispensable for neurodevelopment. Beyond this period, the ablation of even both aPKCs is characterized by a conspicuous absence of anticipated drastic effects. The genetic models developed might prove useful for further interrogating aPKC function in neurodevelopment and neuronal function or to reveal the role of polarity complex function in neurons, astrocytes and oligodendrocytes during stress, injuries or diseases.

developmental biology↗

bioRxiv2024

Time-resolved function of cell polarity kinases PRKCZ and PRKCI in CNS myelination

The atypical Protein Kinase C (aPKC) is a critical component of the PAR polarity complex, as well as of cell polarity in general. There are two paralogs of aPKC in the mammalian genome. Their expression pattern and functional role in oligodendrocyte lineage differentiation and development remains heretofore uncharacterized. Here we show that both paralogs of aPKC - PRKCI, the primary transcript of Prkci and PKMz, an alternate transcript of Prkcz - are expressed during oligodendrocyte differentiation, yet are functionally non-redundant. Asymmetric cell division and early specification of the oligodendrocyte fate stage required both paralogs. By contrast, the stage of oligodendrocyte maturation involving membrane polarization and myelination of axons continued to require PKMz, but no longer needed PRKCI. Taken together, our results indicate to a two-stage role of aPKCs and cell polarity in oligodendrocyte differentiation and development.

developmental biology↗

bioRxiv2024

TAM receptors control actomyosin dynamics in osteoclasts via RHOA-COFILIN-MYOSIN II signaling

The TAM family of receptor tyrosine kinases were recently identified to regulate bone homeostasis by controlling osteoblasts and bone formation. Despite extensive knowledge of TAM receptor function in the mononuclear phagocyte system, the role of TAM receptors in osteoclasts remains largely unknown. Here, we identify a physiological regulatory system including MERTK and TYRO3 in osteoclasts controlling RHOA-ROCK-COFILIN/Myosin II signaling, thereby antagonistically regulating osteoclast-mediated bone remodeling to maintain bone homeostasis. Myeloid-specific lysozyme M-mediated deletion of Mertk led to increased bone mass in mice. In early stages of RANKL-induced osteoclast differentiation MERTK promotes amoeboid migration mode in osteoclast precursor cells by inducing RHOA-COFILIN-MLC2 pathway leading to increased osteoclast formation. In contrast, TYRO3 inhibits RHOA-ROCK signaling in osteoclast precursor cells thereby inhibiting these processes. Furthermore, we unraveled an inhibitory role of MERTK as well as TYRO3 in osteoclast differentiation and function. In line with this, mice with cathepsin K-mediated deletion of Mertk and Tyro3 exhibited an osteoporotic bone phenotype. We found that osteoclast precursor cell morphology dictates its fusion capacity and identified MERTK as a negative regulator of osteoclast fusion. In multinucleated cells, deletion of Mertk inhibits actin ring formation by mediating central actomyosin contraction and inactivation of COFILIN. In contrast, spatially well-ordered RHOA activation at adhesion structures, induced by loss of Tyro3, improves osteoclast biomechanotransduction to ameliorate podosome belt formation and enhance osteoclast function. By using a syngeneic breast cancer bone metastasis osteolysis models we identified TYRO3 as a bone protective receptor for osteolytic bone diseases, whereas MERTK represents a pharmacologic accessible target to inhibit osteoclast formation because its stimulatory effects of osteoclast precursors prevail the inhibitory effects on mature osteoclasts. Next to the recently uncovered role of MERTK as a target for osteoanabolic therapy MERTK may represent a one-drug two-target treatment strategy to increase osteoblast function and reduce osteoclast formation for treatments of bone diseases.

cell biology↗

bioRxiv2023

Stem cells tightly regulate dead cell clearance to maintain tissue fitness

Summary ParagraphMacrophages and dendritic cells have long been appreciated for their ability to migrate to and engulf dying cells and debris, including some of the billions of cells that are naturally eliminated from our body daily. However, a substantial number of these dying cells are cleared by non-professional phagocytes, local epithelial cells that are critical to organismal fitness. How non-professional phagocytes sense and digest nearby apoptotic corpses while still performing their normal tissue functions is unclear. Here, we explore the molecular mechanisms underlying their multifunctionality. Exploiting the cyclical bouts of tissue regeneration and degeneration during the hair cycle, we show that stem cells can transiently become non-professional phagocytes when confronted with dying cells. Adoption of this phagocytic state requires both local lipids produced by apoptotic corpses to activate RXR, and tissue-specific retinoids for RAR{gamma} activation. This dual factor dependency enables tight regulation of the genes requisite to activate phagocytic apoptotic clearance. The tunable phagocytic program we describe here offers an effective mechanism to offset phagocytic duties against the primary stem cell function of replenishing differentiated cells to preserve tissue integrity during homeostasis. Our findings have broad implications for other non-motile stem or progenitor cells which experience cell death in an immune-privileged niche.

cell biology↗

bioRxiv2022

Inflammation Of The Retinal Pigment Epithelium Drives Early-Onset Photoreceptor Degeneration In Mertk-Associated Retinitis Pigmentosa

Severe, early-onset photoreceptor (PR) degeneration associated with MERTK mutations is thought to result from failed phagocytosis by retinal pigment epithelium (RPE). Notwithstanding, the severity and onset of PR degeneration in mouse models of Mertk ablation is determined by the hypomorphic expression or the loss of the Mertk paralog Tyro3. Here we find that loss of Mertk and reduced expression/loss of Tyro3 led to RPE inflammation even before eye-opening. Incipient RPE inflammation cascaded to involve microglia activation and PR degeneration with monocyte infiltration. Inhibition of RPE inflammation with the JAK1/2 inhibitor ruxolitinib mitigated PR degeneration in Mertk-/- mice. Neither inflammation nor severe, early-onset PR degeneration were observed in mice with defective phagocytosis alone. Thus, inflammation drives severe, early-onset PR degeneration-associated with Mertk loss of function.

immunology↗

bioRxiv2022

Tissue-specific modifier alleles determine Mertk loss-of-function traits

Knockout (KO) mouse models play critical roles in elucidating biological processes behind disease-associated or disease-resistant traits. As a consequence of gene KO, mice display certain phenotypes. Based on insight into the molecular role of said gene in a biological process, it is inferred that the particular biological process causally underlies the trait. This approach has been crucial towards understanding the basis of pathological and/or advantageous traits associated with Mertk KO. MERTK is a receptor tyrosine kinase with a critical role in phagocytosis of apoptotic cells or cellular debris. Therefore, early-onset, severe retinal degeneration was described to be a direct consequence of failed phagocytosis of photoreceptor outer segments by retinal pigment epithelia. Similarly, enhanced anti-tumor immunity was inferred to result from the failure of macrophages to dispose cancer cell corpses, resulting in a pro-inflammatory tumor microenvironment. Here we report that the loss of Mertk alone is not sufficient for retinal degeneration. This trait only manifests when the function of the paralog Tyro3 is concomitantly lost. Additionally, the dramatic resistance against two syngeneic mouse tumor models observed in Mertk KO cannot, at least entirely, be ascribed to the loss of Mertk. The widely used Mertk KO carries multiple coincidental changes in its genome that affect the expression of a number of genes, including Tyro3. Nonetheless, neither Tyro3, nor macrophage phagocytosis by alternate genetic redundancy, accounts for the absence of anti-tumor immunity in two independent Mertk KOs. Collectively, our results indicate that context-dependent epistasis of independent modifier alleles determine Mertk KO traits.

immunology↗

bioRxiv2021

Gas6 drives Zika virus-induced neurological complications in humans and congenital syndrome in immunocompetent mice

Zika virus (ZIKV) has the ability to cross placental and brain barriers, causing congenital malformations in neonates and neurological disorders in adults. However, the pathogenic mechanisms of ZIKV-induced neurological complications in adults and congenital malformations remain unknown. Gas6 is a soluble TAM receptor ligand able to promote flavivirus internalization and downregulation of immune responses. Here we demonstrate high Gas6 levels in the serum of patients with neurological complications which correlated with downregulation of genes associated with the type I IFN responses as consequence of Socs1 upregulation. Gas6 gamma-carboxylation is essential for ZIKV replication in monocytes, the main source of this protein. Gas6 also facilitates ZIKV replication in adult immunocompetent mice enabled susceptibility to transplacental infection and congenital malformations. Our data thus indicate that ZIKV promotes the upregulation of its ligand Gas6, which contributes to viral infectivity and drives the development of severe adverse outcomes during ZIKV infection.

microbiology↗

bioRxiv2020

IL-4-induced hysteresis in naive T cell activation

Naive T cells are generally considered to be a homogeneous population, but for their unique T cell receptors (TCRs). Naive T cells are activated within a specific cytokine milieu upon interaction with antigen-presenting cells through cognate TCR::MHC-peptide interaction and co-stimulation. Here we demonstrate that naive T cells are transcriptionally heterogeneous, and that the relative proportions of transcriptionally distinct naive T cell subpopulations are modified by immune responses, such as during helminth infection. Not only are cognate naive T cells activated during an immune response, but the cytokine produced - such as IL-4 during helminth infection - changes the transcriptome of bystander naive T cells. Such changes in gene expression and population level heterogeneity in bystander naive T cells result in altered responses to a concurrent immune challenge, for instance, hypo-responsiveness to vaccination. Thus, naive T cell activation is not the result of a singular temporal event, but is characterized by hysteresis. Our studies suggest that antigen-agnostic, cytokine-dependent naive T cell conditioning and resulting hysteresis is a mechanism that integrates input signals from concurrent infections for the regulation of the overall magnitude of the immune response.

immunology↗