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Interleukin-6 is an activator of pituitary stem cells upon local damage, a competence quenched in the aging gland

Proceedings of the National Academy of Sciences of the United States of America

2021 Jun 22

Vennekens, A;Laporte, E;Hermans, F;Cox, B;Modave, E;Janiszewski, A;Nys, C;Kobayashi, H;Malengier-Devlies, B;Chappell, J;Matthys, P;Garcia, MI;Pasque, V;Lambrechts, D;Vankelecom, H;
PMID: 34161279 | DOI: 10.1073/pnas.2100052118

Stem cells in the adult pituitary are quiescent yet show acute activation upon tissue injury. The molecular mechanisms underlying this reaction are completely unknown. We applied single-cell transcriptomics to start unraveling the acute pituitary stem cell activation process as occurring upon targeted endocrine cell-ablation damage. This stem cell reaction was contrasted with the aging (middle-aged) pituitary, known to have lost damage-repair capacity. Stem cells in the aging pituitary show regressed proliferative activation upon injury and diminished in vitro organoid formation. Single-cell RNA sequencing uncovered interleukin-6 (IL-6) as being up-regulated upon damage, however only in young but not aging pituitary. Administering IL-6 to young mice promptly triggered pituitary stem cell proliferation, while blocking IL-6 or associated signaling pathways inhibited such reaction to damage. By contrast, IL-6 did not generate a pituitary stem cell activation response in aging mice, coinciding with elevated basal IL-6 levels and raised inflammatory state in the aging gland (inflammaging). Intriguingly, in vitro stem cell activation by IL-6 was discerned in organoid culture not only from young but also from aging pituitary, indicating that the aging gland's stem cells retain intrinsic activatability in vivo, likely impeded by the prevailing inflammatory tissue milieu. Importantly, IL-6 supplementation strongly enhanced the growth capability of pituitary stem cell organoids, thereby expanding their potential as an experimental model. Our study identifies IL-6 as a pituitary stem cell activator upon local damage, a competence quenched at aging, concomitant with raised IL-6/inflammatory levels in the older gland. These insights may open the way to interfering with pituitary aging.

Anterior thalamic dysfunction underlies cognitive deficits in a subset of neuropsychiatric disease models

Neuron

2021 Jun 22

Roy, DS;Zhang, Y;Aida, T;Choi, S;Chen, Q;Hou, Y;Lea, NE;Skaggs, KM;Quay, JC;Liew, M;Maisano, H;Le, V;Jones, C;Xu, J;Kong, D;Sullivan, HA;Saunders, A;McCarroll, SA;Wickersham, IR;Feng, G;
PMID: 34197733 | DOI: 10.1016/j.neuron.2021.06.005

Neuropsychiatric disorders are often accompanied by cognitive impairments/intellectual disability (ID). It is not clear whether there are converging mechanisms underlying these debilitating impairments. We found that many autism and schizophrenia risk genes are expressed in the anterodorsal subdivision (AD) of anterior thalamic nuclei, which has reciprocal connectivity with learning and memory structures. CRISPR-Cas9 knockdown of multiple risk genes selectively in AD thalamus led to memory deficits. While the AD is necessary for contextual memory encoding, the neighboring anteroventral subdivision (AV) regulates memory specificity. These distinct functions of AD and AV are mediated through their projections to retrosplenial cortex, using differential mechanisms. Furthermore, knockdown of autism and schizophrenia risk genes PTCHD1, YWHAG, or HERC1 from AD led to neuronal hyperexcitability, and normalization of hyperexcitability rescued memory deficits in these models. This study identifies converging cellular to circuit mechanisms underlying cognitive deficits in a subset of neuropsychiatric disease models.

Early role for a Na+,K+-ATPase (ATP1A3) in brain development

Proceedings of the National Academy of Sciences of the United States of America

2021 Jun 22

Smith, RS;Florio, M;Akula, SK;Neil, JE;Wang, Y;Hill, RS;Goldman, M;Mullally, CD;Reed, N;Bello-Espinosa, L;Flores-Sarnat, L;Monteiro, FP;Erasmo, CB;Pinto E Vairo, F;Morava, E;Barkovich, AJ;Gonzalez-Heydrich, J;Brownstein, CA;McCarroll, SA;Walsh, CA;
PMID: 34161264 | DOI: 10.1073/pnas.2023333118

Osmotic equilibrium and membrane potential in animal cells depend on concentration gradients of sodium (Na+) and potassium (K+) ions across the plasma membrane, a function catalyzed by the Na+,K+-ATPase α-subunit. Here, we describe ATP1A3 variants encoding dysfunctional α3-subunits in children affected by polymicrogyria, a developmental malformation of the cerebral cortex characterized by abnormal folding and laminar organization. To gain cell-biological insights into the spatiotemporal dynamics of prenatal ATP1A3 expression, we built an ATP1A3 transcriptional atlas of fetal cortical development using mRNA in situ hybridization and transcriptomic profiling of ∼125,000 individual cells with single-cell RNA sequencing (Drop-seq) from 11 areas of the midgestational human neocortex. We found that fetal expression of ATP1A3 is most abundant to a subset of excitatory neurons carrying transcriptional signatures of the developing subplate, yet also maintains expression in nonneuronal cell populations. Moving forward a year in human development, we profiled ∼52,000 nuclei from four areas of an infant neocortex and show that ATP1A3 expression persists throughout early postnatal development, most predominantly in inhibitory neurons, including parvalbumin interneurons in the frontal cortex. Finally, we discovered the heteromeric Na+,K+-ATPase pump complex may form nonredundant cell-type-specific α-β isoform combinations, including α3-β1 in excitatory neurons and α3-β2 in inhibitory neurons. Together, the developmental malformation phenotype of affected individuals and single-cell ATP1A3 expression patterns point to a key role for α3 in human cortex development, as well as a cell-type basis for pre- and postnatal ATP1A3-associated diseases.

BMPR-2 gates activity-dependent stabilization of primary dendrites during mitral cell remodeling

Cell reports

2021 Jun 22

Aihara, S;Fujimoto, S;Sakaguchi, R;Imai, T;
PMID: 34161760 | DOI: 10.1016/j.celrep.2021.109276

Developing neurons initially form excessive neurites and then remodel them based on molecular cues and neuronal activity. Developing mitral cells in the olfactory bulb initially extend multiple primary dendrites. They then stabilize single primary dendrites while eliminating others. However, the mechanisms underlying selective dendrite remodeling remain elusive. Using CRISPR-Cas9-based knockout screening combined with in utero electroporation, we identify BMPR-2 as a key regulator for selective dendrite stabilization. Bmpr2 knockout and its rescue experiments show that BMPR-2 inhibits LIMK without ligands and thereby permits dendrite destabilization. In contrast, the overexpression of antagonists and agonists indicates that ligand-bound BMPR-2 stabilizes dendrites, most likely by releasing LIMK. Using genetic and FRET imaging experiments, we demonstrate that free LIMK is activated by NMDARs via Rac1, facilitating dendrite stabilization through F-actin formation. Thus, the selective stabilization of primary dendrites is ensured by concomitant inputs of BMP ligands and neuronal activity.

miRNA Changes in Retinal Ganglion Cells after Optic Nerve Crush and Glaucomatous Damage

Cells

2021 Jun 22

Mead, B;Kerr, A;Nakaya, N;Tomarev, SI;
PMID: 34206213 | DOI: 10.3390/cells10071564

The purpose of this study was to characterize the miRNA profile of purified retinal ganglion cells (RGC) from healthy and diseased rat retina. Diseased retina includes those after a traumatic optic nerve crush (ONC), and after ocular hypertension/glaucoma. Rats were separated into four groups: healthy/intact, 7 days after laser-induced ocular hypertension, 2 days after traumatic ONC, and 7 days after ONC. RGC were purified from rat retina using microbeads conjugated to CD90.1/Thy1. RNA were sequenced using Next Generation Sequencing. Over 100 miRNA were identified that were significantly different in diseased retina compared to healthy retina. Considerable differences were seen in the miRNA expression of RGC 7 days after ONC, whereas after 2 days, few changes were seen. The miRNA profiles of RGC 7 days after ONC and 7 days after ocular hypertension were similar, but discrete miRNA differences were still seen. Candidate mRNA showing different levels of expression after retinal injury were manipulated in RGC cultures using mimics/AntagomiRs. Of the five candidate miRNA identified and subsequently tested for therapeutic efficacy, miR-194 inhibitor and miR-664-2 inhibitor elicited significant RGC neuroprotection, whereas miR-181a mimic and miR-181d-5p mimic elicited significant RGC neuritogenesis.

Humans with inherited T cell CD28 deficiency are susceptible to skin papillomaviruses but are otherwise healthy

Cell

2021 Jun 22

Béziat, V;Rapaport, F;Hu, J;Titeux, M;Bonnet des Claustres, M;Bourgey, M;Griffin, H;Bandet, É;Ma, CS;Sherkat, R;Rokni-Zadeh, H;Louis, DM;Changi-Ashtiani, M;Delmonte, OM;Fukushima, T;Habib, T;Guennoun, A;Khan, T;Bender, N;Rahman, M;About, F;Yang, R;Rao, G;Rouzaud, C;Li, J;Shearer, D;Balogh, K;Al Ali, F;Ata, M;Dabiri, S;Momenilandi, M;Nammour, J;Alyanakian, MA;Leruez-Ville, M;Guenat, D;Materna, M;Marcot, L;Vladikine, N;Soret, C;Vahidnezhad, H;Youssefian, L;Saeidian, AH;Uitto, J;Catherinot, É;Navabi, SS;Zarhrate, M;Woodley, DT;Jeljeli, M;Abraham, T;Belkaya, S;Lorenzo, L;Rosain, J;Bayat, M;Lanternier, F;Lortholary, O;Zakavi, F;Gros, P;Orth, G;Abel, L;Prétet, JL;Fraitag, S;Jouanguy, E;Davis, MM;Tangye, SG;Notarangelo, LD;Marr, N;Waterboer, T;Langlais, D;Doorbar, J;Hovnanian, A;Christensen, N;Bossuyt, X;Shahrooei, M;Casanova, JL;
PMID: 34214472 | DOI: 10.1016/j.cell.2021.06.004

We study a patient with the human papilloma virus (HPV)-2-driven "tree-man" phenotype and two relatives with unusually severe HPV4-driven warts. The giant horns form an HPV-2-driven multifocal benign epithelial tumor overexpressing viral oncogenes in the epidermis basal layer. The patients are unexpectedly homozygous for a private CD28 variant. They have no detectable CD28 on their T cells, with the exception of a small contingent of revertant memory CD4+ T cells. T cell development is barely affected, and T cells respond to CD3 and CD2, but not CD28, costimulation. Although the patients do not display HPV-2- and HPV-4-reactive CD4+ T cells in vitro, they make antibodies specific for both viruses in vivo. CD28-deficient mice are susceptible to cutaneous infections with the mouse papillomavirus MmuPV1. The control of HPV-2 and HPV-4 in keratinocytes is dependent on the T cell CD28 co-activation pathway. Surprisingly, human CD28-dependent T cell responses are largely redundant for protective immunity.

Endogenous reverse transcriptase and RNase H-mediated antiviral mechanism in embryonic stem cells

Cell research

2021 Jun 22

Wu, J;Wu, C;Xing, F;Cao, L;Zeng, W;Guo, L;Li, P;Zhong, Y;Jiang, H;Luo, M;Shi, G;Bu, L;Ji, Y;Hou, P;Peng, H;Huang, J;Li, C;Guo, D;
PMID: 34158624 | DOI: 10.1038/s41422-021-00524-7

Nucleic acid-based systems play important roles in antiviral defense, including CRISPR/Cas that adopts RNA-guided DNA cleavage to prevent DNA phage infection and RNA interference (RNAi) that employs RNA-guided RNA cleavage to defend against RNA virus infection. Here, we report a novel type of nucleic acid-based antiviral system that exists in mouse embryonic stem cells (mESCs), which suppresses RNA virus infection by DNA-mediated RNA cleavage. We found that the viral RNA of encephalomyocarditis virus can be reverse transcribed into complementary DNA (vcDNA) by the reverse transcriptase (RTase) encoded by endogenous retrovirus-like elements in mESCs. The vcDNA is negative-sense single-stranded and forms DNA/RNA hybrid with viral RNA. The viral RNA in the heteroduplex is subsequently destroyed by cellular RNase H1, leading to robust suppression of viral growth. Furthermore, either inhibition of the RTase activity or depletion of endogenous RNase H1 results in the promotion of virus proliferation. Altogether, our results provide intriguing insights into the antiviral mechanism of mESCs and the antiviral function of endogenized retroviruses and cellular RNase H. Such a natural nucleic acid-based antiviral mechanism in mESCs is referred to as ERASE (endogenous RTase/RNase H-mediated antiviral system), which is an addition to the previously known nucleic acid-based antiviral mechanisms including CRISPR/Cas in bacteria and RNAi in plants and invertebrates.

Developmental and sexual dimorphic atlas of the prenatal mouse external genitalia at the single-cell level

Proceedings of the National Academy of Sciences of the United States of America

2021 Jun 22

Amato, CM;Yao, HH;
PMID: 34155146 | DOI: 10.1073/pnas.2103856118

Birth defects of the external genitalia are among the most common in the world. Proper formation of the external genitalia requires a highly orchestrated process that involves special cell populations and sexually dimorphic hormone signaling. It is clear what the end result of the sexually dimorphic development is (a penis in the male versus clitoris in the female); however, the cell populations involved in the process remain poorly defined. Here, we used single-cell messenger RNA sequencing in mouse embryos to uncover the dynamic changes in cell populations in the external genitalia during the critical morphogenetic window. We found that overall, male and female external genitalia are largely composed of the same core cellular components. At the bipotential stage of development (embryonic day or E14.5), few differences in cell populational composition exist between male and female. Although similar in cell population composition, genetic differences in key sexual differentiation developmental pathways arise between males and females by the early (E16.5) and late (E18.5) differentiation stages. These differences include discrete cell populations with distinct responsiveness to androgen and estrogen. By late sexual differentiation (E18.5), unique cell populations in both male and female genitalia become apparent and are enriched with androgen- and estrogen-responsive genes, respectively. These data provide insights into the morphogenesis of the external genitalia that could be used to understand diseases associated with defects in the external genitalia.

Protein arginine methyltransferase 1 regulates cell proliferation and differentiation in adult mouse adult intestine

Cell & bioscience

2021 Jun 22

Xue, L;Bao, L;Roediger, J;Su, Y;Shi, B;Shi, YB;
PMID: 34158114 | DOI: 10.1186/s13578-021-00627-z

Adult stem cells play an essential role in adult organ physiology and tissue repair and regeneration. While much has been learnt about the property and function of various adult stem cells, the mechanisms of their development remain poorly understood in mammals. Earlier studies suggest that the formation of adult mouse intestinal stem cells takes place during the first few weeks after birth, the postembryonic period when plasma thyroid hormone (T3) levels are high. Furthermore, deficiency in T3 signaling leads to defects in adult mouse intestine, including reduced cell proliferation in the intestinal crypts, where stem cells reside. Our earlier studies have shown that protein arginine methyltransferase 1 (PRMT1), a T3 receptor coactivator, is highly expressed during intestinal maturation in mouse.We have analyzed the expression of PRMT1 by immunohistochemistry and studied the effect of tissue-specific knockout of PRMT1 in the intestinal epithelium.We show that PRMT1 is expressed highly in the proliferating transit amplifying cells and crypt base stem cells. By using a conditional knockout mouse line, we have demonstrated that the expression of PRMT1 in the intestinal epithelium is critical for the development of the adult mouse intestine. Specific removal of PRMT1 in the intestinal epithelium results in, surprisingly, more elongated adult intestinal crypts with increased cell proliferation. In addition, epithelial cell migration along the crypt-villus axis and cell death on the villus are also increased. Furthermore, there are increased Goblet cells and reduced Paneth cells in the crypt while the number of crypt base stem cells remains unchanged.Our finding that PRMT1 knockout increases cell proliferation is surprising considering the role of PRMT1 in T3-signaling and the importance of T3 for intestinal development, and suggests that PRMT1 likely regulates pathways in addition to T3-signaling to affect intestinal development and/or homeostasis, thus affecting cell proliferating and epithelial turn over in the adult.

Estrogen receptor-α signaling in post-natal mammary development and breast cancers

Cellular and molecular life sciences : CMLS

2021 Jun 22

Rusidzé, M;Adlanmérini, M;Chantalat, E;Raymond-Letron, I;Cayre, S;Arnal, JF;Deugnier, MA;Lenfant, F;
PMID: 34156490 | DOI: 10.1007/s00018-021-03860-4

17β-estradiol controls post-natal mammary gland development and exerts its effects through Estrogen Receptor ERα, a member of the nuclear receptor family. ERα is also critical for breast cancer progression and remains a central therapeutic target for hormone-dependent breast cancers. In this review, we summarize the current understanding of the complex ERα signaling pathways that involve either classical nuclear "genomic" or membrane "non-genomic" actions and regulate in concert with other hormones the different stages of mammary development. We describe the cellular and molecular features of the luminal cell lineage expressing ERα and provide an overview of the transgenic mouse models impacting ERα signaling, highlighting the pivotal role of ERα in mammary gland morphogenesis and function and its implication in the tumorigenic processes. Finally, we describe the main features of the ERα-positive luminal breast cancers and their modeling in mice.

Distinct populations of antigen specific tissue resident CD8 T cells in human cervix mucosa

JCI insight

2021 Jun 22

Peng, T;Phasouk, K;Bossard, E;Klock, A;Jin, L;Laing, KJ;Johnston, C;Williams, NA;Czartoski, JL;Varon, D;Long, AN;Bielas, JH;Snyder, TM;Robins, H;Koelle, DM;McElrath, MJ;Wald, A;Corey, L;Zhu, J;
PMID: 34156975 | DOI: 10.1172/jci.insight.149950

The ectocervix is part of the lower female reproductive tract (FRT), which is susceptible to sexually transmitted infections (STI). Comprehensive knowledge of the phenotypes and T cell receptor (TCR) repertoire of tissue resident memory T cells (TRM) in human FRT is lacking. We have taken single-cell RNA sequencing approaches to simultaneously define gene expression and TCR clonotypes of the human ectocervix. There are significantly more CD8 than CD4 T cells. Unsupervised clustering and trajectory analysis identify distinct populations of CD8 T cells with IFNGhiGZMBlowCD69hiCD103low or IFNGlowGZMBhiCD69medCD103hi phenotypes. Little overlap was seen between their TCR repertoires. Immunofluorescent staining shows that CD103+ CD8 TRM cells preferentially localize in epithelium while CD69+ CD8 TRM distribute evenly in epithelium and stroma. Ex vivo assays indicate up to 14% of cervical CD8 TRM clonotypes are HSV-2 reactive in HSV-2-seropositive persons, reflecting physiologically relevant localization. Our studies identify subgroups of CD8 TRM in the human ectocervix that exhibit distinct expression of antiviral defense and tissue residency markers, anatomic locations, and TCR repertoires that target anatomically relevant viral antigens. Optimization of the location, number, and function of FRT TRM is an important approach for improving host defenses to STI.

Castration-induced down-regulation of SPARC in stromal cells drives neuroendocrine differentiation of prostate cancer

Cancer research

2021 Jun 21

Enriquez, C;Cancila, V;Ferri, R;Sulsenti, R;Fischetti, I;Milani, M;Ostano, P;Gregnanin, I;Mello-Grand, M;Berrino, E;Bregni, M;Renne, G;Tripodo, C;Colombo, MP;Jachetti, E;
PMID: 34185677 | DOI: 10.1158/0008-5472.CAN-21-0163

Fatal neuroendocrine differentiation (NED) of castration-resistant prostate cancer is a recurrent mechanism of resistance to androgen deprivation therapies (ADT) and anti-androgen receptor pathway inhibitors (ARPI) in patients. The design of effective therapies for neuroendocrine prostate cancer (NEPC) is complicated by limited knowledge of the molecular mechanisms governing NED. The paucity of acquired genomic alterations and the deregulation of epigenetic and transcription factors suggest a potential contribution from the microenvironment. In this context, whether ADT/ARPI induces stromal cells to release NED-promoting molecules and the underlying molecular networks are unestablished. Here, we utilized transgenic and transplantable mouse models and co-culture experiments to unveil a novel tumor-stroma crosstalk that is able to induce NED under the pressure of androgen deprivation. Castration induced upregulation of GRP78 in tumor cells, which triggers miR-29b-mediated downregulation of the matricellular protein SPARC in the nearby stroma. SPARC downregulation enabled stromal cells to release IL-6, a known inducer of NED. A drug that targets GRP78 blocked NED in castrated mice. A public, human NEPC gene expression dataset showed that Hspa5 (encoding for GRP78) positively correlates with hallmarks of NED. Finally, prostate cancer specimens from patients developing local NED after ADT showed GRP78 upregulation in tumor cells and SPARC downregulation in the stroma. These results point to GRP78 as a potential therapeutic target and to SPARC downregulation in stromal cells as a potential early biomarker of tumors undergoing NED.

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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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