Dataset Information

Secretome analysis of C-mannosyltransferase-deficient hiPSCs

ABSTRACT: C-mannosylation is a modification of tryptophan residues with a single mannose effecting protein folding, secretion and/or function. To date, only few proteins have been proven to be C-mannosylated and studies aiming at global assessment of protein C-mannosylation from cells or tissues are scarce. In order to interrogate the C-mannosylome of human induced pluripotent stem cells (hiPSCs), we made use of the common finding that C-mannosylation is important for protein secretion. Thus we compared the secretomes of C-mannosyltransferase-deficient DPY19L1 and DPY19L3 mutants to their parental wild-type hiPSCs by mass-spectrometry-based quantitative proteomics. Secretion of numerous proteins was reduced in the mutants.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stem Cell, Cell Culture

SUBMITTER: Karsten Cirksena

LAB HEAD: Falk Büttner

PROVIDER: PXD024193 | Pride | 2021-06-17

REPOSITORIES: Pride

ACCESS DATA

Publications

The C-Mannosylome of Human Induced Pluripotent Stem Cells Implies a Role for ADAMTS16 C-Mannosylation in Eye Development.

Cirksena Karsten K Hütte Hermann J HJ Shcherbakova Aleksandra A Thumberger Thomas T Sakson Roman R Weiss Stefan S Jensen Lars Riff LR Friedrich Alina A Todt Daniel D Kuss Andreas W AW Ruppert Thomas T Wittbrodt Joachim J Bakker Hans H Buettner Falk F R FFR

Molecular & cellular proteomics : MCP 20210508

C-mannosylation is a modification of tryptophan residues with a single mannose and can affect protein folding, secretion, and/or function. To date, only a few proteins have been demonstrated to be C-mannosylated, and studies that globally assess protein C-mannosylation are scarce. To interrogate the C-mannosylome of human induced pluripotent stem cells, we compared the secretomes of CRISPR-Cas9 mutants lacking either the C-mannosyltransferase DPY19L1 or DPY19L3 to WT human induced pluripotent st ...[more]

PMID: 33975020

Similar Datasets

Project description:ABSTRACT Stimulating the commitment of implanted dystrophin+ muscle derived stem cells (MDSC) into myogenic, as opposed to lipofibrogenic, lineages is a promising therapeutic strategy for Duchenne muscular dystrophy (DMD). To examine whether counteracting myostatin, a negative regulator of muscle mass and a pro-lipofibrotic factor, would help this process, we compared the in vitro myogenic and fibrogenic capacity of MDSC from wild type (WT), myostatin knockout (Mst KO), and mdx (DMD model) (mdx) young mice under various modulators, the expression of key stem cell and myogenic genes, and the capacity of these MDSC to repair the injured gastrocnemius in aged mdx mice with exacerbated lipofibrosis. Surprisingly, the potent in vitro myotube formation by WT MDSC was refractory to modulators of myostatin expression or activity, and the Mst KO and mdx MDSC failed to form myotubes under any condition, despite all MDSC expressed Oct-4 and various stem cell genes and differentiated into other lineages. The genetic inactivation of myostatin or dystrophin in MDSC was associated with silencing of critical genes for early myogenesis (Actc1, Acta1, and MyoD). WT MDSC implanted into the injured gastrocnemius of old mdx mice significantly improved myofiber repair and reduced fat deposition and, to a lesser extent, fibrosis. In contrast to their in vitro behavior, Mst KO MDSC in vivo also significantly improved myofiber repair, but had no significant effects on lipofibrotic degeneration. In conclusion, while WT MDSC are considerably myogenic in culture and stimulate muscle repair after injury in the aged mdx mouse, myostatin genetic inactivation blocks myotube formation in vitro but the myogenic capacity is recovered in vivo under the influence of the host tissue environment, presumably by reactivation of key genes originally silenced in the Mst KO MDSC. Key words: dystrophin, mdx mouse, Duchenne, fibrosis, dystrophy One sample each of mouse wild type (WT), myostatin knockout (KO), muscular dystrophy model mdx, and the transgenic OCT4 promoter plus reporter were grown, RNA was isolated, and subjected to the SABiosciences mouse stem cell oligo array.

Project description:The two-component system CBO0366/CBO0365 was recently demonstrated to have a role in cold tolerance of Group I Clostridium botulinum ATCC 3502. The mechanisms under its control, ultimately resulting in increased sensitivity to low temperature, are unknown. A transcriptomic analysis with DNA microarrays was performed to identify the differences in global gene expression patterns of the wild-type ATCC 3502 and a derivative mutant with insertionally inactivated cbo0365 at 37 M-BM-0C and 15 M-BM-0C. Altogether 150 or 141 chromosomal CDSs were found to be differently expressed in the cbo0365 mutant at 37 M-BM-0C or 15 M-BM-0C, respectively, and thus considered to be under direct or indirect transcriptional control of the response regulator CBO0365. Of the differentially-expressed CDSs, expression of 141 CDSs was similarly affected at both temperatures investigated, suggesting that the putative CBO0365 regulon was practically not affected by temperature. The regulon involved genes related to acetone-butanol-ethanol (ABE) fermentation, motility, to arsenic resistance, and phosphate uptake and transport. Deteriorated growth at 17 M-BM-0C was observed for mutants with disrupted ABE fermentation pathway components (crt, bdh and ctfA), arsenic detoxifying machinery components (arsC and arsR), or phosphate uptake mechanism components (phoT), suggesting roles for these mechanisms in cold tolerance of Group I C. botulinum. Electrophoretic mobility shift assays showed recombinant CBO0365 to bind to the promoter regions of crt, arsR, and phoT, as well as to the promoter region of its own operon, suggesting direct DNA-binding transcriptional activation or repression as means for CBO0365 in regulating these operons. The results provide insight to the mechanisms Group I C. botulinum utilize in coping with cold. C. botulinum ATCC 3502 cbo0365 mutant vs. wild type; 3 replicates of each strain; growth at 37C in TPGY broth batch culture and subjected to cold shock to 15C; sampling at mid-log growth phase before cold shock, and 1 h after temperature downshift to 15C (= 2 time points). Dye-swapped hybridization.

Dataset Information

Secretome analysis of C-mannosyltransferase-deficient hiPSCs

Publications

The C-Mannosylome of Human Induced Pluripotent Stem Cells Implies a Role for ADAMTS16 C-Mannosylation in Eye Development.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure