DNAJB1-PRKACA in HEK293T cells induces LINC00473 overexpression that depends on PKA signaling
Fibrolamellar carcinoma (FLC) is a primary liver cancer that most commonly arises in adolescents and young adults in a background of normal liver tissue and has a poor prognosis due to lack of effective chemotherapeutic agents. The DNAJB1-PRKACA gene fusion (DP) has been reported in the majority of...
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| Veröffentlicht in: | PloS one 2022-02, Vol.17 (2), p.e0263829-e0263829 |
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| creator | Kim, Stephanie S Kycia, Ina Karski, Michael Ma, Rosanna K Bordt, Evan A Kwan, Julian Karki, Anju Winter, Elle Aktas, Ranan G Wu, Yuxuan Emili, Andrew Bauer, Daniel E Sethupathy, Praveen Vakili, Khashayar |
| description | Fibrolamellar carcinoma (FLC) is a primary liver cancer that most commonly arises in adolescents and young adults in a background of normal liver tissue and has a poor prognosis due to lack of effective chemotherapeutic agents. The DNAJB1-PRKACA gene fusion (DP) has been reported in the majority of FLC tumors; however, its oncogenic mechanisms remain unclear. Given the paucity of cellular models, in particular FLC tumor cell lines, we hypothesized that engineering the DP fusion gene in HEK293T cells would provide insight into the cellular effects of the fusion gene. We used CRISPR/Cas9 to engineer HEK293T clones expressing DP fusion gene (HEK-DP) and performed transcriptomic, proteomic, and mitochondrial studies to characterize this cellular model. Proteomic analysis of DP interacting partners identified mitochondrial proteins as well as proteins in other subcellular compartments. HEK-DP cells demonstrated significantly elevated mitochondrial fission, which suggests a role for DP in altering mitochondrial dynamics. Transcriptomic analysis of HEK-DP cells revealed a significant increase in LINC00473 expression, similar to what has been observed in primary FLC samples. LINC00473 overexpression was reversible with siRNA targeting of PRKACA as well as pharmacologic targeting of PKA and Hsp40 in HEK-DP cells. Therefore, our model suggests that LINC00473 is a candidate marker for DP activity. |
| doi_str_mv | 10.1371/journal.pone.0263829 |
| format | Article |
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The DNAJB1-PRKACA gene fusion (DP) has been reported in the majority of FLC tumors; however, its oncogenic mechanisms remain unclear. Given the paucity of cellular models, in particular FLC tumor cell lines, we hypothesized that engineering the DP fusion gene in HEK293T cells would provide insight into the cellular effects of the fusion gene. We used CRISPR/Cas9 to engineer HEK293T clones expressing DP fusion gene (HEK-DP) and performed transcriptomic, proteomic, and mitochondrial studies to characterize this cellular model. Proteomic analysis of DP interacting partners identified mitochondrial proteins as well as proteins in other subcellular compartments. HEK-DP cells demonstrated significantly elevated mitochondrial fission, which suggests a role for DP in altering mitochondrial dynamics. Transcriptomic analysis of HEK-DP cells revealed a significant increase in LINC00473 expression, similar to what has been observed in primary FLC samples. LINC00473 overexpression was reversible with siRNA targeting of PRKACA as well as pharmacologic targeting of PKA and Hsp40 in HEK-DP cells. Therefore, our model suggests that LINC00473 is a candidate marker for DP activity.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0263829</identifier><identifier>PMID: 35167623</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Adolescents ; Biochemistry ; Biology and Life Sciences ; Carcinoma, Hepatocellular - genetics ; Cell fusion ; Cell lines ; Chemotherapy ; Cloning ; CRISPR ; CRISPR-Cas Systems ; Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - genetics ; Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - metabolism ; Engineering ; Engineering and Technology ; Fusion protein ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Gene fusion ; Genetic aspects ; Health aspects ; HEK293 Cells ; Hematology ; Hospitals ; HSP40 Heat-Shock Proteins - genetics ; HSP40 Heat-Shock Proteins - metabolism ; Hsp40 protein ; Humans ; Kinases ; Liver ; Liver cancer ; Medical prognosis ; Medicine and Health Sciences ; Mitochondria ; Mitochondria - metabolism ; Models, Biological ; Oncogene Proteins, Fusion - genetics ; Oncology ; Pediatrics ; Protein kinase A ; Protein kinases ; Proteins ; Proteomics ; Research and Analysis Methods ; RNA ; RNA, Long Noncoding - genetics ; siRNA ; Stem cells ; Surgery ; Transcriptomics ; Tumor cell lines ; Tumors ; Up-Regulation ; Veterinary colleges ; Veterinary medicine ; Young adults</subject><ispartof>PloS one, 2022-02, Vol.17 (2), p.e0263829-e0263829</ispartof><rights>COPYRIGHT 2022 Public Library of Science</rights><rights>2022 Kim et al. This is an open access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2022 Kim et al 2022 Kim et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c6079-a8474872ad9a889e1c6c5ebbfdd318a05827dc652c2e6335d3007ca6df724ee73</citedby><cites>FETCH-LOGICAL-c6079-a8474872ad9a889e1c6c5ebbfdd318a05827dc652c2e6335d3007ca6df724ee73</cites><orcidid>0000-0003-2542-2237</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC8846505/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC8846505/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,864,885,2102,2928,23866,27924,27925,53791,53793,79600,79601</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/35167623$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Roemer, Klaus</contributor><creatorcontrib>Kim, Stephanie S</creatorcontrib><creatorcontrib>Kycia, Ina</creatorcontrib><creatorcontrib>Karski, Michael</creatorcontrib><creatorcontrib>Ma, Rosanna K</creatorcontrib><creatorcontrib>Bordt, Evan A</creatorcontrib><creatorcontrib>Kwan, Julian</creatorcontrib><creatorcontrib>Karki, Anju</creatorcontrib><creatorcontrib>Winter, Elle</creatorcontrib><creatorcontrib>Aktas, Ranan G</creatorcontrib><creatorcontrib>Wu, Yuxuan</creatorcontrib><creatorcontrib>Emili, Andrew</creatorcontrib><creatorcontrib>Bauer, Daniel E</creatorcontrib><creatorcontrib>Sethupathy, Praveen</creatorcontrib><creatorcontrib>Vakili, Khashayar</creatorcontrib><title>DNAJB1-PRKACA in HEK293T cells induces LINC00473 overexpression that depends on PKA signaling</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Fibrolamellar carcinoma (FLC) is a primary liver cancer that most commonly arises in adolescents and young adults in a background of normal liver tissue and has a poor prognosis due to lack of effective chemotherapeutic agents. The DNAJB1-PRKACA gene fusion (DP) has been reported in the majority of FLC tumors; however, its oncogenic mechanisms remain unclear. Given the paucity of cellular models, in particular FLC tumor cell lines, we hypothesized that engineering the DP fusion gene in HEK293T cells would provide insight into the cellular effects of the fusion gene. We used CRISPR/Cas9 to engineer HEK293T clones expressing DP fusion gene (HEK-DP) and performed transcriptomic, proteomic, and mitochondrial studies to characterize this cellular model. Proteomic analysis of DP interacting partners identified mitochondrial proteins as well as proteins in other subcellular compartments. HEK-DP cells demonstrated significantly elevated mitochondrial fission, which suggests a role for DP in altering mitochondrial dynamics. Transcriptomic analysis of HEK-DP cells revealed a significant increase in LINC00473 expression, similar to what has been observed in primary FLC samples. LINC00473 overexpression was reversible with siRNA targeting of PRKACA as well as pharmacologic targeting of PKA and Hsp40 in HEK-DP cells. Therefore, our model suggests that LINC00473 is a candidate marker for DP activity.</description><subject>Adolescents</subject><subject>Biochemistry</subject><subject>Biology and Life Sciences</subject><subject>Carcinoma, Hepatocellular - genetics</subject><subject>Cell fusion</subject><subject>Cell lines</subject><subject>Chemotherapy</subject><subject>Cloning</subject><subject>CRISPR</subject><subject>CRISPR-Cas Systems</subject><subject>Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - genetics</subject><subject>Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - metabolism</subject><subject>Engineering</subject><subject>Engineering and Technology</subject><subject>Fusion protein</subject><subject>Gene Expression Profiling</subject><subject>Gene Expression Regulation, Neoplastic</subject><subject>Gene fusion</subject><subject>Genetic aspects</subject><subject>Health aspects</subject><subject>HEK293 Cells</subject><subject>Hematology</subject><subject>Hospitals</subject><subject>HSP40 Heat-Shock Proteins - genetics</subject><subject>HSP40 Heat-Shock Proteins - metabolism</subject><subject>Hsp40 protein</subject><subject>Humans</subject><subject>Kinases</subject><subject>Liver</subject><subject>Liver cancer</subject><subject>Medical prognosis</subject><subject>Medicine and Health Sciences</subject><subject>Mitochondria</subject><subject>Mitochondria - metabolism</subject><subject>Models, Biological</subject><subject>Oncogene Proteins, Fusion - genetics</subject><subject>Oncology</subject><subject>Pediatrics</subject><subject>Protein kinase A</subject><subject>Protein kinases</subject><subject>Proteins</subject><subject>Proteomics</subject><subject>Research and Analysis Methods</subject><subject>RNA</subject><subject>RNA, Long Noncoding - genetics</subject><subject>siRNA</subject><subject>Stem cells</subject><subject>Surgery</subject><subject>Transcriptomics</subject><subject>Tumor cell lines</subject><subject>Tumors</subject><subject>Up-Regulation</subject><subject>Veterinary colleges</subject><subject>Veterinary medicine</subject><subject>Young 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AMP-Dependent Protein Kinase Catalytic Subunits - metabolism</topic><topic>Engineering</topic><topic>Engineering and Technology</topic><topic>Fusion protein</topic><topic>Gene Expression Profiling</topic><topic>Gene Expression Regulation, Neoplastic</topic><topic>Gene fusion</topic><topic>Genetic aspects</topic><topic>Health aspects</topic><topic>HEK293 Cells</topic><topic>Hematology</topic><topic>Hospitals</topic><topic>HSP40 Heat-Shock Proteins - genetics</topic><topic>HSP40 Heat-Shock Proteins - metabolism</topic><topic>Hsp40 protein</topic><topic>Humans</topic><topic>Kinases</topic><topic>Liver</topic><topic>Liver cancer</topic><topic>Medical prognosis</topic><topic>Medicine and Health Sciences</topic><topic>Mitochondria</topic><topic>Mitochondria - metabolism</topic><topic>Models, Biological</topic><topic>Oncogene Proteins, Fusion - genetics</topic><topic>Oncology</topic><topic>Pediatrics</topic><topic>Protein kinase A</topic><topic>Protein kinases</topic><topic>Proteins</topic><topic>Proteomics</topic><topic>Research and Analysis Methods</topic><topic>RNA</topic><topic>RNA, Long Noncoding - genetics</topic><topic>siRNA</topic><topic>Stem cells</topic><topic>Surgery</topic><topic>Transcriptomics</topic><topic>Tumor cell lines</topic><topic>Tumors</topic><topic>Up-Regulation</topic><topic>Veterinary colleges</topic><topic>Veterinary medicine</topic><topic>Young adults</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kim, Stephanie S</creatorcontrib><creatorcontrib>Kycia, Ina</creatorcontrib><creatorcontrib>Karski, Michael</creatorcontrib><creatorcontrib>Ma, Rosanna K</creatorcontrib><creatorcontrib>Bordt, Evan A</creatorcontrib><creatorcontrib>Kwan, Julian</creatorcontrib><creatorcontrib>Karki, Anju</creatorcontrib><creatorcontrib>Winter, Elle</creatorcontrib><creatorcontrib>Aktas, Ranan G</creatorcontrib><creatorcontrib>Wu, Yuxuan</creatorcontrib><creatorcontrib>Emili, 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A</au><au>Kwan, Julian</au><au>Karki, Anju</au><au>Winter, Elle</au><au>Aktas, Ranan G</au><au>Wu, Yuxuan</au><au>Emili, Andrew</au><au>Bauer, Daniel E</au><au>Sethupathy, Praveen</au><au>Vakili, Khashayar</au><au>Roemer, Klaus</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>DNAJB1-PRKACA in HEK293T cells induces LINC00473 overexpression that depends on PKA signaling</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2022-02-15</date><risdate>2022</risdate><volume>17</volume><issue>2</issue><spage>e0263829</spage><epage>e0263829</epage><pages>e0263829-e0263829</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Fibrolamellar carcinoma (FLC) is a primary liver cancer that most commonly arises in adolescents and young adults in a background of normal liver tissue and has a poor prognosis due to lack of effective chemotherapeutic agents. The DNAJB1-PRKACA gene fusion (DP) has been reported in the majority of FLC tumors; however, its oncogenic mechanisms remain unclear. Given the paucity of cellular models, in particular FLC tumor cell lines, we hypothesized that engineering the DP fusion gene in HEK293T cells would provide insight into the cellular effects of the fusion gene. We used CRISPR/Cas9 to engineer HEK293T clones expressing DP fusion gene (HEK-DP) and performed transcriptomic, proteomic, and mitochondrial studies to characterize this cellular model. Proteomic analysis of DP interacting partners identified mitochondrial proteins as well as proteins in other subcellular compartments. HEK-DP cells demonstrated significantly elevated mitochondrial fission, which suggests a role for DP in altering mitochondrial dynamics. Transcriptomic analysis of HEK-DP cells revealed a significant increase in LINC00473 expression, similar to what has been observed in primary FLC samples. LINC00473 overexpression was reversible with siRNA targeting of PRKACA as well as pharmacologic targeting of PKA and Hsp40 in HEK-DP cells. Therefore, our model suggests that LINC00473 is a candidate marker for DP activity.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>35167623</pmid><doi>10.1371/journal.pone.0263829</doi><tpages>e0263829</tpages><orcidid>https://orcid.org/0000-0003-2542-2237</orcidid><oa>free_for_read</oa></addata></record> |
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| issn | 1932-6203 1932-6203 |
| language | eng |
| recordid | cdi_plos_journals_2629049346 |
| source | MEDLINE; DOAJ Directory of Open Access Journals; Public Library of Science (PLoS) Journals Open Access; EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Adolescents Biochemistry Biology and Life Sciences Carcinoma, Hepatocellular - genetics Cell fusion Cell lines Chemotherapy Cloning CRISPR CRISPR-Cas Systems Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - genetics Cyclic AMP-Dependent Protein Kinase Catalytic Subunits - metabolism Engineering Engineering and Technology Fusion protein Gene Expression Profiling Gene Expression Regulation, Neoplastic Gene fusion Genetic aspects Health aspects HEK293 Cells Hematology Hospitals HSP40 Heat-Shock Proteins - genetics HSP40 Heat-Shock Proteins - metabolism Hsp40 protein Humans Kinases Liver Liver cancer Medical prognosis Medicine and Health Sciences Mitochondria Mitochondria - metabolism Models, Biological Oncogene Proteins, Fusion - genetics Oncology Pediatrics Protein kinase A Protein kinases Proteins Proteomics Research and Analysis Methods RNA RNA, Long Noncoding - genetics siRNA Stem cells Surgery Transcriptomics Tumor cell lines Tumors Up-Regulation Veterinary colleges Veterinary medicine Young adults |
| title | DNAJB1-PRKACA in HEK293T cells induces LINC00473 overexpression that depends on PKA signaling |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-02T14%3A02%3A36IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_plos_&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=DNAJB1-PRKACA%20in%20HEK293T%20cells%20induces%20LINC00473%20overexpression%20that%20depends%20on%20PKA%20signaling&rft.jtitle=PloS%20one&rft.au=Kim,%20Stephanie%20S&rft.date=2022-02-15&rft.volume=17&rft.issue=2&rft.spage=e0263829&rft.epage=e0263829&rft.pages=e0263829-e0263829&rft.issn=1932-6203&rft.eissn=1932-6203&rft_id=info:doi/10.1371/journal.pone.0263829&rft_dat=%3Cgale_plos_%3EA693704374%3C/gale_plos_%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=2629049346&rft_id=info:pmid/35167623&rft_galeid=A693704374&rft_doaj_id=oai_doaj_org_article_932a91da3a8749ee88d4d191f4c126d6&rfr_iscdi=true |