Extracellular mechanical forces drive endocardial cell volume decrease during zebrafish cardiac valve morphogenesis

Developmental cell

2022 Mar 14

Vignes, H;Vagena-Pantoula, C;Prakash, M;Fukui, H;Norden, C;Mochizuki, N;Jug, F;Vermot, J;
PMID: 35245444 | DOI: 10.1016/j.devcel.2022.02.011

Organ morphogenesis involves dynamic changes of tissue properties while cells adapt to their mechanical environment through mechanosensitive pathways. How mechanical cues influence cell behaviors during morphogenesis remains unclear. Here, we studied the formation of the zebrafish atrioventricular canal (AVC) where cardiac valves develop. We show that the AVC forms within a zone of tissue convergence associated with the increased activation of the actomyosin meshwork and cell-orientation changes. We demonstrate that tissue convergence occurs with a reduction of cell volume triggered by mechanical forces and the mechanosensitive channel TRPP2/TRPV4. Finally, we show that the extracellular matrix component hyaluronic acid controls cell volume changes. Together, our data suggest that multiple force-sensitive signaling pathways converge to modulate cell volume. We conclude that cell volume reduction is a key cellular feature activated by mechanotransduction during cardiovascular morphogenesis. This work further identifies how mechanical forces and extracellular matrix influence tissue remodeling in developing organs.

The spatial transcriptomic landscape of the healing mouse intestine following damage

Nature communications

2022 Feb 11

Parigi, SM;Larsson, L;Das, S;Ramirez Flores, RO;Frede, A;Tripathi, KP;Diaz, OE;Selin, K;Morales, RA;Luo, X;Monasterio, G;Engblom, C;Gagliani, N;Saez-Rodriguez, J;Lundeberg, J;Villablanca, EJ;
PMID: 35149721 | DOI: 10.1038/s41467-022-28497-0

The intestinal barrier is composed of a complex cell network defining highly compartmentalized and specialized structures. Here, we use spatial transcriptomics to define how the transcriptomic landscape is spatially organized in the steady state and healing murine colon. At steady state conditions, we demonstrate a previously unappreciated molecular regionalization of the colon, which dramatically changes during mucosal healing. Here, we identified spatially-organized transcriptional programs defining compartmentalized mucosal healing, and regions with dominant wired pathways. Furthermore, we showed that decreased p53 activation defined areas with increased presence of proliferating epithelial stem cells. Finally, we mapped transcriptomics modules associated with human diseases demonstrating the translational potential of our dataset. Overall, we provide a publicly available resource defining principles of transcriptomic regionalization of the colon during mucosal healing and a framework to develop and progress further hypotheses.

SRSF1 governs progenitor-specific alternative splicing to maintain adult epithelial tissue homeostasis and renewal

Developmental cell

2022 Feb 22

Yu, T;Cazares, O;Tang, AD;Kim, HY;Wald, T;Verma, A;Liu, Q;Barcellos-Hoff, MH;Floor, SN;Jung, HS;Brooks, AN;Klein, OD;
PMID: 35202586 | DOI: 10.1016/j.devcel.2022.01.011

Alternative splicing generates distinct mRNA variants and is essential for development, homeostasis, and renewal. Proteins of the serine/arginine (SR)-rich splicing factor family are major splicing regulators that are broadly required for organ development as well as cell and organism viability. However, how these proteins support adult organ function remains largely unknown. Here, we used the continuously growing mouse incisor as a model to dissect the functions of the prototypical SR family protein SRSF1 during tissue homeostasis and renewal. We identified an SRSF1-governed alternative splicing network that is specifically required for dental proliferation and survival of progenitors but dispensable for the viability of differentiated cells. We also observed a similar progenitor-specific role of SRSF1 in the small intestinal epithelium, indicating a conserved function of SRSF1 across adult epithelial tissues. Thus, our findings define a regulatory mechanism by which SRSF1 specifically controls progenitor-specific alternative splicing events to support adult tissue homeostasis and renewal.

Structure-Based Design of a Chemical Probe Set for the 5-HT5A Serotonin Receptor

Journal of medicinal chemistry

2022 Feb 23

Levit Kaplan, A;Strachan, RT;Braz, JM;Craik, V;Slocum, S;Mangano, T;Amabo, V;O'Donnell, H;Lak, P;Basbaum, AI;Roth, BL;Shoichet, BK;
PMID: 35195401 | DOI: 10.1021/acs.jmedchem.1c02031

The 5-HT5A receptor (5-HT5AR), for which no selective agonists and a few antagonists exist, remains the least understood serotonin receptor. A single commercial antagonist, SB-699551, has been widely used to investigate the 5-HT5AR function in neurological disorders, including pain, but this molecule has substantial liabilities as a chemical probe. Accordingly, we sought to develop an internally controlled probe set. Docking over 6 million molecules against a 5-HT5AR homology model identified 5 mid-μM ligands, one of which was optimized to UCSF678, a 42 nM arrestin-biased partial agonist at the 5-HT5AR with a more restricted off-target profile and decreased assay liabilities versus SB-699551. Site-directed mutagenesis supported the docked pose of UCSF678. Surprisingly, analogs of UCSF678 that lost the 5-HT5AR activity revealed that 5-HT5AR engagement is nonessential for alleviating pain, contrary to studies with less-selective ligands. UCSF678 and analogs constitute a selective probe set with which to study the function of the 5-HT5AR.

Towards Tabula Gallus

International journal of molecular sciences

2022 Jan 06

Yamagata, M;
PMID: 35054796 | DOI: 10.3390/ijms23020613

The Tabula Gallus is a proposed project that aims to create a map of every cell type in the chicken body and chick embryos. Chickens (Gallus gallus) are one of the most recognized model animals that recapitulate the development and physiology of mammals. The Tabula Gallus will generate a compendium of single-cell transcriptome data from Gallus gallus, characterize each cell type, and provide tools for the study of the biology of this species, similar to other ongoing cell atlas projects (Tabula Muris and Tabula Sapiens/Human Cell Atlas for mice and humans, respectively). The Tabula Gallus will potentially become an international collaboration between many researchers. This project will be useful for the basic scientific study of Gallus gallus and other birds (e.g., cell biology, molecular biology, developmental biology, neuroscience, physiology, oncology, virology, behavior, ecology, and evolution). It will eventually be beneficial for a better understanding of human health and diseases.

Single-cell reconstruction with spatial context of migrating neural crest cells and their microenvironments during vertebrate head and neck formation

Development

2021 Nov 15

Morrison, J;McLennan, R;Teddy, J;Scott, A;Kasemeier-Kulesa, J;Gogol, M;Kulesa, P;
| DOI: 10.1242/dev.199468

The dynamics of multipotent neural crest cell differentiation and invasion as cells travel throughout the vertebrate embryo remain unclear. Here, we preserve spatial information to derive the transcriptional states of migrating neural crest cells and the cellular landscape of the first four chick cranial to cardiac branchial arches (BA1-4) using label-free, unsorted single-cell RNA sequencing. The faithful capture of branchial arch-specific genes led to identification of novel markers of migrating neural crest cells and 266 invasion genes common to all BA1-4 streams. Perturbation analysis of a small subset of invasion genes and time-lapse imaging identified their functional role to regulate neural crest cell behaviors. Comparison of the neural crest invasion signature to other cell invasion phenomena revealed a shared set of 45 genes, a subset of which showed direct relevance to human neuroblastoma cell lines analyzed after exposure to the in vivo chick embryonic neural crest microenvironment. Our data define an important spatio-temporal reference resource to address patterning of the vertebrate head and neck, and previously unidentified cell invasion genes with the potential for broad impact.

A proteogenomic portrait of lung squamous cell carcinoma

Cell

2021 Aug 05

Satpathy, S;Krug, K;Jean Beltran, PM;Savage, SR;Petralia, F;Kumar-Sinha, C;Dou, Y;Reva, B;Kane, MH;Avanessian, SC;Vasaikar, SV;Krek, A;Lei, JT;Jaehnig, EJ;Omelchenko, T;Geffen, Y;Bergstrom, EJ;Stathias, V;Christianson, KE;Heiman, DI;Cieslik, MP;Cao, S;Song, X;Ji, J;Liu, W;Li, K;Wen, B;Li, Y;Gümüş, ZH;Selvan, ME;Soundararajan, R;Visal, TH;Raso, MG;Parra, ER;Babur, Ö;Vats, P;Anand, S;Schraink, T;Cornwell, M;Rodrigues, FM;Zhu, H;Mo, CK;Zhang, Y;da Veiga Leprevost, F;Huang, C;Chinnaiyan, AM;Wyczalkowski, MA;Omenn, GS;Newton, CJ;Schurer, S;Ruggles, KV;Fenyö, D;Jewell, SD;Thiagarajan, M;Mesri, M;Rodriguez, H;Mani, SA;Udeshi, ND;Getz, G;Suh, J;Li, QK;Hostetter, G;Paik, PK;Dhanasekaran, SM;Govindan, R;Ding, L;Robles, AI;Clauser, KR;Nesvizhskii, AI;Wang, P;Carr, SA;Zhang, B;Mani, DR;Gillette, MA;Clinical Proteomic Tumor Analysis Consortium, ;
PMID: 34358469 | DOI: 10.1016/j.cell.2021.07.016

Lung squamous cell carcinoma (LSCC) remains a leading cause of cancer death with few therapeutic options. We characterized the proteogenomic landscape of LSCC, providing a deeper exposition of LSCC biology with potential therapeutic implications. We identify NSD3 as an alternative driver in FGFR1-amplified tumors and low-p63 tumors overexpressing the therapeutic target survivin. SOX2 is considered undruggable, but our analyses provide rationale for exploring chromatin modifiers such as LSD1 and EZH2 to target SOX2-overexpressing tumors. Our data support complex regulation of metabolic pathways by crosstalk between post-translational modifications including ubiquitylation. Numerous immune-related proteogenomic observations suggest directions for further investigation. Proteogenomic dissection of CDKN2A mutations argue for more nuanced assessment of RB1 protein expression and phosphorylation before declaring CDK4/6 inhibition unsuccessful. Finally, triangulation between LSCC, LUAD, and HNSCC identified both unique and common therapeutic vulnerabilities. These observations and proteogenomics data resources may guide research into the biology and treatment of LSCC.

Epithelial STAT6 O-GlcNAcylation Drives Anti-Helminth Immunity via a Concerted Alarmin Response

SSRN Electronic Journal

2021 Sep 08

Zhao, M;Ren, K;Xiong, X;Xin, Y;Kim, A;Maynard, J;Zou, Y;Battist, A;Koneripalli, N;Huang, Z;Zhang, Z;Yu, J;Wang, H;Salgado, O;Hogquist, K;Revelo, X;Burlingame, A;Gao, X;Lin, Z;von Moltke, J;Ruan, H;
| DOI: 10.2139/ssrn.3917158

The epithelium is an integral component of mucosal barrier and host immunity. Following helminth parasite infection, the intestinal epithelial cells secrete "alarmin” cytokines, such as interleukin (IL)-25 and IL-33, to initiate the type 2 immune responses for helminth expulsion and tolerance. However, it is unknown how helminth infection and the resulting type 2 cytokine milieu drive epithelial remodeling and orchestrate alarmin secretion. Here we report that, intestinal epithelial O-linked N-Acetylglucosamine (O-GlcNAc) protein modification is induced upon helminth infections. By modifying and activating STAT6, O-GlcNAc transferase (OGT) promotes the transcription of lineage-defining transcription factor _Pou2f3_ in tuft cell differentiation and IL-25 production. Meanwhile, STAT6 O-GlcNAcylation activates the expression of _Gsdmc_ family genes. The resulting membrane pore formed by GSDMC facilitates the unconventional secretion of IL-33 from goblet cells. GSDMC-mediated IL-33 secretion is indispensable for the host to mount effective antihelminth immunity and support intestinal homeostasis. Protein O-GlcNAcylation can be harnessed for the future treatment of type 2 inflammation-associated human diseases.

Avances en citometría de masas y aplicabilidad en patología digital para estudios clínico-traslacionales en oncología

Advances in Laboratory Medicine / Avances en Medicina de Laboratorio

2021 Aug 25

Cereceda, K;Jorquera, R;Villarroel-Espíndola, F;
| DOI: 10.1515/almed-2021-0051

Resumen El desarrollo de la citometría de masas y posteriormente su adaptación para el análisis de secciones histológicas ha revolucionado la forma de caracterizar a nivel espacial múltiples componentes de manera simultánea, permitiendo la correlación genotípica y fenotípica de la célula y su entorno durante estudios clínicos-traslaciones. En este trabajo, hemos revisado los hitos más relevantes en el desarrollo, implementación y aplicabilidad del análisis de imágenes de componentes múltiples para el estudio de cáncer y otras dolencias, y enfocado nuestro interés que aquellos autores que utilizan imágenes obtenidas mediante citometría de masas o bien haz de iones. Esta revisión tiene como objetivo que el lector se familiarice con las estrategias técnicas de verificación de la herramienta y las múltiples posibilidades de uso abordadas por diferentes autores, y además, poder proyectar sus propias investigaciones hacia la utilización de imágenes obtenidas por citometría de masas (IMC), o imágenes por haz de iones multiplexados (MIBI) en cualquiera de los campos de investigación biomédica.

Prognostic value and cost benefit of HPV testing for oropharyngeal cancer patients

Oral diseases

2021 Jun 15

Lu, XJD;Liu, KYP;Prisman, E;Wu, J;Zhu, YS;Poh, C;
PMID: 34129700 | DOI: 10.1111/odi.13938

High-risk human papillomavirus (HR-HPV) can cause oropharyngeal squamous cell carcinoma (OpSCC). The revised 8th edition of the AJCC Staging Manual now stages OpSCC by incorporating p16 immunohistochemistry (IHC), the surrogate marker for HPV status. This study assessed the prognostic values of p16 and HPV markers.We identified 244 OpSCC patients diagnosed between 2000-2008 from the British Columbia Cancer Registry with enough tissue to conduct experiments. Formalin-fixed, paraffin-embedded tissue sections were stained for p16 IHC, RNA in situ hybridization (ISH) HPV 16 and 18, and DNA ISH HR-HPV. Electronic charts were reviewed to collect clinical and outcome data. Combined positive RNA and/or DNA ISH was used to denote HPV status.HPV was positive among 77.9% of samples. Using HPV as the benchmark, p16 IHC had high sensitivity (90.5%), but low specificity (68.5%). Distinct subgroups of patients were identified by sequential separation of p16 then HPV status. Among both p16-positive and p16-negative groups, HPV-positive patients were younger, more males, and had better clinical outcomes, especially 5-year overall survival. We further evaluated the technical costs associated with HPV testing.HPV is more prognostic than p16 for OpSCC. Clinical laboratories can adopt HPV RNA ISH for routine analysis.This article is protected by

Hypoxia-induced lncHILAR promotes renal cancer cell invasion and metastasis via ceRNA for the miR-613/206/1-1-3p/Jagged-1/Notch/CXCR4 signaling pathway

Molecular therapy : the journal of the American Society of Gene Therapy

2021 May 28

Hu, G;Ma, J;Zhang, J;Chen, Y;Liu, H;Huang, Y;Zheng, J;Xu, Y;Xue, W;Zhai, W;
PMID: 34058384 | DOI: 10.1016/j.ymthe.2021.05.020

Hypoxia has been identified as a common driving factor that contributes to tumor progression, including invasion and metastasis. However, the underlying mechanisms of enhanced invasion and metastasis under hypoxia remain unclear. A hypoxic microenvironment promoted invasion and metastasis of RCC by upregulating the expression of LOC100506178, which we named Hypoxia-Induced lncRNA Associated with Renal Cell Carcinoma (lncHILAR). Knockdown of lncHILAR inhibited cell invasion and migration while overexpression of lncHILAR conversely facilitated cell invasion and migration of RCC cells. Notably, hypoxic RCC cells secreted exosomes packaged with lncHILAR which were taken up by normoxic RCC cells and then drove normoxic cell invasion. Mechanistically, hypoxia-induced-lncHILAR elevated RCC invasion and metastasis by acting as a competing endogenous (ce)RNA for miR-613/206/1-1-3p, which led to the upregulation of Jagged-1 and C-X-C Motif Chemokine Receptor 4 (CXCR4). Activation of the of Jagged-1/Notch/CXCR4 axis induced RCC metastasis. Hypoxia-induced lncHILAR promotes RCC cell invasion and metastasis via ceRNA for the miR-613/206/1-1-3p/Jagged-1/Notch/CXCR4 axis. The novel lncHILAR may thus serve as a potential biomarker and therapeutic target in RCC.

Lateral ventral tegmental area GABAergic and glutamatergic modulation of conditioned learning

Cell reports

2021 Mar 16

Rizzi, G;Li, Z;Hogrefe, N;Tan, KR;
PMID: 33730568 | DOI: 10.1016/j.celrep.2021.108867

The firing activity of dorso-medial-striatal-cholinergic interneurons (dmCINs) is a neural correlate of classical conditioning. Tonically active, they pause in response to salient stimuli, mediating acquisition of predictive cues/outcome associations. Cortical and thalamic inputs are typical of the rather limited knowledge about underlying circuitry contributing to this function. Here, we dissect the midbrain GABA and glutamate-to-dmCIN pathways and evaluate how they influence conditioned behavior. We report that midbrain neurons discriminate auditory cues and encode the association of a predictive stimulus with a footshock. Furthermore, GABA and glutamate cells form selective monosynaptic contacts onto dmCINs and di-synaptic ones via the parafascicular thalamus. Pathway-specific inhibition of each sub-circuit produces differential impairments of fear-conditioned learning. Finally, Vglut2-expressing cells discriminate between CSs although Vgat-positive neurons associate the predictive cue with the outcome. Overall, these data suggest that each component of the network carries information pertinent to sub-domains of the behavioral strategy.

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	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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