LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk
Although papillary thyroid carcinoma (PTC) has a good prognosis, 20–90% of patients show metastasis to regional lymph nodes and 10–15% of patients show metastasis to distant sites. Metastatic disease represents the main clinical challenge that impacts survival rate. We previously showed that LIMD2 w...
| Main Authors: | , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2020-11-01
|
| Series: | Cells |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2073-4409/9/11/2522 |
| _version_ | 1827701706550411264 |
|---|---|
| author | Rodrigo Pinheiro Araldi Thatiana Correa de Melo Débora Levy Dener Madeiro de Souza Beatriz Maurício Gabriel Avelar Colozza-Gama Sergio Paulo Bydlowski Hongzhuang Peng Frank J. Rauscher Janete Maria Cerutti |
| author_facet | Rodrigo Pinheiro Araldi Thatiana Correa de Melo Débora Levy Dener Madeiro de Souza Beatriz Maurício Gabriel Avelar Colozza-Gama Sergio Paulo Bydlowski Hongzhuang Peng Frank J. Rauscher Janete Maria Cerutti |
| author_sort | Rodrigo Pinheiro Araldi |
| collection | DOAJ |
| description | Although papillary thyroid carcinoma (PTC) has a good prognosis, 20–90% of patients show metastasis to regional lymph nodes and 10–15% of patients show metastasis to distant sites. Metastatic disease represents the main clinical challenge that impacts survival rate. We previously showed that LIMD2 was a novel metastasis-associated gene. In this study, to interrogate the role of LIMD2 in cancer invasion and metastasis, we used CRISPR-mediated knockout (KO) of LIMD2 in PTC cells (BCPAP and TPC1). Western blot and high-content screening (HCS) analysis confirmed functional KO of LIMD2. LIMD2 KO reduced in vitro invasion and migration. Ultrastructural analyses showed that cell polarity and mitochondria function and morphology were restored in LIMD2 KO cells. To unveil the signals supervising these phenotypic changes, we employed phospho-protein array. Several members of the MAPK superfamily showed robust reduction in phosphorylation. A Venn diagram displayed the overlap of kinases with reduced phosphorylation in both cell lines and showed that they were able to initiate or sustain the epithelial-mesenchymal transition (EMT) and DNA damage checkpoint. Flow cytometry and HCS validation analyses further corroborated the phospho-protein array data. Collectively, our findings show that LIMD2 enhances phosphorylation of kinases associated with EMT and invasion. Through cooperation with different kinases, it contributes to the increased genomic instability that ultimately promotes PTC progression. |
| first_indexed | 2024-03-10T14:39:09Z |
| format | Article |
| id | doaj.art-a2d37603262541e6a54de04780fa2753 |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-10T14:39:09Z |
| publishDate | 2020-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-a2d37603262541e6a54de04780fa27532023-11-20T21:56:26ZengMDPI AGCells2073-44092020-11-01911252210.3390/cells9112522LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK CrosstalkRodrigo Pinheiro Araldi0Thatiana Correa de Melo1Débora Levy2Dener Madeiro de Souza3Beatriz Maurício4Gabriel Avelar Colozza-Gama5Sergio Paulo Bydlowski6Hongzhuang Peng7Frank J. Rauscher8Janete Maria Cerutti9Genetic Bases of Thyroid Tumors Laboratory, Division of Genetics, Department of Morphology and Genetics, Universidade Federal de São Paulo/EPM, São Paulo, SP 04039-032, BrazilPrograma de Pós-graduação em Biociências, Universidade Federal da Integração Latino-Americana (UNILA), Foz do Iguaçu, PR 85866-000, BrazilLaboratory of Histocompatibility and Cellular Immunity, Faculdade de Medicina, Universidade de São Paulo (USP), São Paulo, SP 05404-000, BrazilGenetics Laboratory, Instituto Butantan, São Paulo, SP 05503-900, BrazilLaboratory of Cell Biology, Instituto Butantan, São Paulo, SP 05503-900, BrazilGenetic Bases of Thyroid Tumors Laboratory, Division of Genetics, Department of Morphology and Genetics, Universidade Federal de São Paulo/EPM, São Paulo, SP 04039-032, BrazilLaboratory of Histocompatibility and Cellular Immunity, Faculdade de Medicina, Universidade de São Paulo (USP), São Paulo, SP 05404-000, BrazilThe Wistar Institute, Philadelphia, PA 19104, USAThe Wistar Institute, Philadelphia, PA 19104, USAGenetic Bases of Thyroid Tumors Laboratory, Division of Genetics, Department of Morphology and Genetics, Universidade Federal de São Paulo/EPM, São Paulo, SP 04039-032, BrazilAlthough papillary thyroid carcinoma (PTC) has a good prognosis, 20–90% of patients show metastasis to regional lymph nodes and 10–15% of patients show metastasis to distant sites. Metastatic disease represents the main clinical challenge that impacts survival rate. We previously showed that LIMD2 was a novel metastasis-associated gene. In this study, to interrogate the role of LIMD2 in cancer invasion and metastasis, we used CRISPR-mediated knockout (KO) of LIMD2 in PTC cells (BCPAP and TPC1). Western blot and high-content screening (HCS) analysis confirmed functional KO of LIMD2. LIMD2 KO reduced in vitro invasion and migration. Ultrastructural analyses showed that cell polarity and mitochondria function and morphology were restored in LIMD2 KO cells. To unveil the signals supervising these phenotypic changes, we employed phospho-protein array. Several members of the MAPK superfamily showed robust reduction in phosphorylation. A Venn diagram displayed the overlap of kinases with reduced phosphorylation in both cell lines and showed that they were able to initiate or sustain the epithelial-mesenchymal transition (EMT) and DNA damage checkpoint. Flow cytometry and HCS validation analyses further corroborated the phospho-protein array data. Collectively, our findings show that LIMD2 enhances phosphorylation of kinases associated with EMT and invasion. Through cooperation with different kinases, it contributes to the increased genomic instability that ultimately promotes PTC progression.https://www.mdpi.com/2073-4409/9/11/2522papillary thyroid carcinomaLIMD2BRAF V600ECRISPR/Cas9epithelial-to-mesenchymal transition |
| spellingShingle | Rodrigo Pinheiro Araldi Thatiana Correa de Melo Débora Levy Dener Madeiro de Souza Beatriz Maurício Gabriel Avelar Colozza-Gama Sergio Paulo Bydlowski Hongzhuang Peng Frank J. Rauscher Janete Maria Cerutti LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk Cells papillary thyroid carcinoma LIMD2 BRAF V600E CRISPR/Cas9 epithelial-to-mesenchymal transition |
| title | LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk |
| title_full | LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk |
| title_fullStr | LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk |
| title_full_unstemmed | LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk |
| title_short | LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk |
| title_sort | limd2 regulates key steps of metastasis cascade in papillary thyroid cancer cells via mapk crosstalk |
| topic | papillary thyroid carcinoma LIMD2 BRAF V600E CRISPR/Cas9 epithelial-to-mesenchymal transition |
| url | https://www.mdpi.com/2073-4409/9/11/2522 |
| work_keys_str_mv | AT rodrigopinheiroaraldi limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT thatianacorreademelo limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT deboralevy limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT denermadeirodesouza limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT beatrizmauricio limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT gabrielavelarcolozzagama limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT sergiopaulobydlowski limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT hongzhuangpeng limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT frankjrauscher limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk AT janetemariacerutti limd2regulateskeystepsofmetastasiscascadeinpapillarythyroidcancercellsviamapkcrosstalk |