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Endothelial cells (ECs) in the brain communicate with mural cells to facilitate vascular stability. Platelet-derived growth factor-BB (PDGF-BB)/platelet-derived growth factor receptor-β (PDGFR-β) signaling mechanism at EC-mural cell interface helps stabilize the vasculature. How this paracrine signaling is mediated is not known. Our laboratory studies endothelial cilia, a microtubule-based organelle, and its role in promoting vascular stability. We discovered that brain endothelial cilia are located primarily on the basolateral side, and PDGF-BB is expressed in EC cilium. Thus, we hypothesized that endothelium cilium in conjunction with PDGF-BB on the basolateral side is responsible for mural cell recruitment. In this study, using a combination of zebrafish, mice, and human brain model systems, we have established a signaling paradigm wherein p21-activated kinase (PAK2) and ADP-ribosylation factor-13b (ARL13b) in ECs induce secretion of PDGF-BB. PDGF-BB associates with heparan sulfate proteoglycans (HSPGs) to form a gradient around ECs. Disrupting PAK2 affects ciliogenesis, HSPGs, and PDGF-BB gradient. We unravel a new mechanism involving endothelial cilia/PAK2-mediated PDGF-BB secretion, and retention by periendothelial HSPGs to promote vascular stability via recruiting mural cells.

The centromere is a part of the chromosome that is essential for the even segregation of duplicated chromosomes during cell division. It is epigenetically defined by the presence of the histone H3 variant CENP-A. CENP-A associates specifically with a group of 16 proteins that form the constitutive centromere-associated network (CCAN) of proteins. In mitosis, the kinetochore forms on the CCAN to connect the duplicated chromosomes to the microtubules protruding from the cell poles. Previous studies have shown that CENP-A replaces H3 in nucleosomes, and recently, the structures of CENP-A-containing nucleosomes in complex with CCANs have been revealed, but they show only a limited interaction between CCANs and CENP-A. Here, we report the cryo-EM structure of 2x(CENP-A/H4)₂ octasomes assembled on DNA in the absence of H2A/H2B histone dimer and speculate how (CENP-A/H4)₂ tetrasomes might serve as a platform for CCAN organization.

GPR88 is an orphan G protein-coupled receptor that regulates dopamine neurotransmission and is a target for neuropsychiatric disorders. In addition to the somatic membrane, GPR88 can localize to the primary cilium, a membrane microdomain known for dynamically enriching receptors and signaling molecules. However, the distribution of GPR88 in neuronal primary cilia remains uncharacterized. Here, we characterize GPR88 distribution at primary cilia in two brain areas. We show that in the striatum, GPR88 localizes both to somatodendritic and primary cilia compartments on inhibitory GABAergic medium spiny neurons. In contrast, in the somatosensory cortex, GPR88 localizes to somatodendritic and nuclear compartments and not primary cilia of excitatory spiny stellate neurons. In addition, we found that cilia density and length were similar between Gpr88 knockout and wild-type animals. Together, we provide key evidence for neuronal cell-type specific regulation of GPR88 localization to primary cilia, suggesting neuron subtype specific regulatory mechanisms govern receptor ciliary targeting in the brain.

T-lymphocyte activation is a crucial process in the regulation of innate and adaptive immune responses. The ion channel-kinase TRPM7, transient receptor potential cation channel subfamily M, member 7, has previously been implicated in cellular Mg²⁺ homeostasis, proliferation, and immune cell modulation. Here, we show that pharmacological and genetic silencing of TRPM7 leads to diminished activation and influences signaling pathways that guide human T_H17 or T_reg cell differentiation, following TCR-mediated stimulation. In primary human CD4 T cells and CRISPR-Cas9-engineered Jurkat T cells, inactivation or loss of TRPM7 led to distorted Mg²⁺ homeostasis and Ca²⁺ signaling, reduced NFAT translocation, decreased IL-2 secretion and altered T_H cell differentiation. While the activation of primary human CD4 T cells, as well as in vitro polarization into pro-inflammatory T_H17 cells was critically dependent on TRPM7, the polarization of naïve CD4 T cells into FOXP3⁺ regulatory T cells was not. Taken together, these results highlight TRPM7 as molecular switch in lymphocyte activation and polarization. Thus, suggesting a therapeutic potential for TRPM7 in numerous T-cell mediated diseases.

Human organotypic brain slice cultures have emerged as a pivotal tool to study the complexities of the human brain. Human organotypic brain slice cultures preserve the structural integrity, cellular diversity, and vascular networks of living brain tissue, maintaining in vivo characteristics. This advancement enables accurate temporal modeling of neurological diseases and facilitates precise experimental manipulations, accelerating therapeutic development. However, their use raises important ethical and philosophical considerations, including issues of donor consent and the potential for neural activity that prompts questions about consciousness. This study outlines these emerging concerns, emphasizing the need for guidelines that balance scientific innovation with ethical responsibility, particularly in relation to donor consent, transparency, and long-term use of living human tissue.

Alternative splicing (AS) and nonsense-mediated mRNA decay (NMD) are highly conserved cellular mechanisms that modulate gene expression. Here, we introduce the NMD pipeline that computes how splicing events introduce premature termination codons to mRNA transcripts via frameshift, then predicts the rate of premature termination codon-dependent NMD. We use whole-blood, deep RNA-sequencing data from critically ill patients to study gene expression in sepsis. Statistical significance was determined as adjusted P < 0.05 and |log₂ fold change| > 2 for differential gene expression and probability ≥0.9 and |DeltaPsi| > 0.1 for AS. The NMD pipeline was developed based on the AS data from Whippet. We demonstrate that the rate of NMD is higher in the sepsis and deceased groups compared with the control and survived groups, which may signify aberrant splicing because of altered physiology in critical illness. Predominance of non-exon skipping events was associated with disease and mortality states. The NMD pipeline also revealed proteins with potential association with sepsis. Together, these results emphasize the utility of the NMD pipeline in studying AS-NMD along with differential gene expression analysis and uncovering proteins associated with sepsis.

Wnt signaling pathways are involved in various developmental and tissue maintenance functions, whereas deregulated Wnt signaling is closely linked to human cancer. Recent work revealed that loss of Wnt signaling impairs mitosis and causes abnormal microtubule growth at the mitotic spindle resulting in chromosome missegregation and aneuploidy, both of which are hallmarks of cancer cells exhibiting chromosomal instability (CIN). Here, we show that upon DNA replication stress, a condition typically associated with CIN, Wnt10b acts to prevent increased microtubule dynamics from the S phase until mitosis, thereby ensuring faithful chromosome segregation. Interestingly, replication stress-induced chromosomal breaks are also efficiently suppressed by Wnt10b. Thus, our results show that Wnt10b signaling regulates replication stress-induced chromosome missegregation and breakage, and hence is a determinant for broad genome instability in cancer cells.

Centriole and/or cilium defects are characteristic of cancer cells and have been linked to cancer cell invasion. However, the mechanistic bases of this regulation remain incompletely understood. Spindle assembly abnormal protein 6 homolog (SAS-6) is essential for centriole biogenesis and cilium formation. SAS-6 levels decrease at the end of mitosis and G1, resulting from APC^Cdh1-targeted degradation. To examine the biological consequences of unrestrained SAS-6 expression, we used a nondegradable SAS-6 mutant (SAS-6ND). This led to an increase in ciliation and cell invasion and caused an up-regulation of the YAP/TAZ pathway. SAS-6ND expression resulted in cell morphology changes, nuclear deformation, and YAP translocation to the nucleus, resulting in increased TEAD-dependent transcription. SAS-6-mediated invasion was prevented by YAP down-regulation or by blocking ciliogenesis. Similarly, down-regulation of SAS-6 in DMS273, a highly invasive and highly ciliated lung cancer cell line that overexpresses SAS-6, completely blocked cell invasion and depleted YAP protein levels. Thus, our data provide evidence for a defined role of SAS-6 in cell invasion through the activation of the YAP/TAZ pathway.

Natural antisense transcripts (AST) are expressed in eukaryotes, prokaryotes, and viruses and can possess regulatory functions at the transcriptional and/or post-transcriptional levels. In vitro studies have shown that HIV-1 AST promote viral latency through epigenetic silencing of the proviral 5' long terminal repeat. However, expression of AST in vivo has not been convincingly demonstrated. Here, we used single RNA template amplification and sequencing to demonstrate expression of AST in unstimulated PBMC collected from people with HIV-1 (PWH). Our results show that expression levels of AST could be higher during ART compared with untreated individuals and that clones of infected cells persisting under ART continue to express HIV AST. This study is the first to verify HIV-1 AST expression in vivo with sequencing, documenting AST presence without cellular activation and suggest its natural occurrence in PWH. These findings advance our understanding of HIV-1 persistence and underscore the need for larger studies to determine if targeting AST in viral reservoirs could lead to new approaches for the design of strategies towards achieving HIV remission without ART.