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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic <t>epithelial</t> cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.
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Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic epithelial cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.

Journal: The Journal of Biological Chemistry

Article Title: Clostridioides difficile TcdB induces expression of its receptor (CSPG4) through a noncanonical Hippo signaling mechanism

doi: 10.1016/j.jbc.2026.111137

Figure Lengend Snippet: Core Hippo kinase regulation of CSPG4 in TcdB exposed pericytes . A , pericytes were exposed to 1 ng/ml of TcdB2 for 24 h and colonic epithelial cells were exposed to 10 ng/ml of TcdB2 for 24 h. Multiple wells from each treatment were combined for RNA isolation. RT-qPCR was used to quantify CSPG4 transcripts. RT-qPCR data are presented as mean (n = 3) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. B , Immunoblots from protein lysates acquired from pericytes exposed for 24 h to 1 ng/ml of TcdB. C , immunoblots from protein lysates obtained from pericytes or HeLa cells exposed for 24 h to 1 ng/ml of TcdB. Phosphorylated LATS1 (pLATS1) was detected with an antibody recognizing phosphorylation at Thr 1079. D , densitometry analysis of pLATS1 immunoblots. Relative band density is presented as mean (n = 2) ± S.D. with each data point as a technical replicate. ∗ p < 0.05 determined by Student’s t test. E , immunoblots from protein lysates acquired from pericyte or HeLa cells exposed for 24 h to 1 ng/ml of TcdB in the presence or absence of 10 μM XMU-MP-1. F , immunoblots from protein lysates taken from HeLa cells exposed for 24 h to 1 ng/ml of TcdB with and without of 30 μM TRULI.

Article Snippet: Primary human colonic epithelial cells (36,037–08) were obtained from Celprogen and were cultured using the protocol provided by Celprogen.

Techniques: Isolation, Quantitative RT-PCR, Western Blot, Phospho-proteomics