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Analysis of NIS Plasma Membrane Interactors Discloses Key Regulation by a SRC/RAC1/PAK1/PIP5K/EZRIN Pathway with Potential Implications for Radioiodine Re-Sensitization Therapy in Thyroid Cancer

2021-10-30Journal article Open access
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dc.contributor.authorFaria, Márcia
dc.contributor.authorDomingues, Rita
dc.contributor.authorBugalho, Maria João
dc.contributor.authorSilva, Ana Luísa
dc.contributor.authorMatos, Paulo
dc.date.accessioned2022-02-03T17:12:19Z
dc.date.available2022-02-03T17:12:19Z
dc.date.issued2021-10-30
dc.descriptionThis article belongs to the Special Issue Role of Small GTPase Signaling in Tumorigenesispt_PT
dc.description.abstractThe functional expression of the sodium-iodide symporter (NIS) at the membrane of differentiated thyroid cancer (DTC) cells is the cornerstone for the use of radioiodine (RAI) therapy in these malignancies. However, NIS gene expression is frequently downregulated in malignant thyroid tissue, and 30% to 50% of metastatic DTCs become refractory to RAI treatment, which dramatically decreases patient survival. Several strategies have been attempted to increase the NIS mRNA levels in refractory DTC cells, so as to re-sensitize refractory tumors to RAI. However, there are many RAI-refractory DTCs in which the NIS mRNA and protein levels are relatively abundant but only reduced levels of iodide uptake are detected, suggesting a posttranslational failure in the delivery of NIS to the plasma membrane (PM), or an impaired residency at the PM. Because little is known about the molecules and pathways regulating NIS delivery to, and residency at, the PM of thyroid cells, we here employed an intact-cell labeling/immunoprecipitation methodology to selectively purify NIS-containing macromolecular complexes from the PM. Using mass spectrometry, we characterized and compared the composition of NIS PM complexes to that of NIS complexes isolated from whole cell (WC) lysates. Applying gene ontology analysis to the obtained MS data, we found that while both the PM-NIS and WC-NIS datasets had in common a considerable number of proteins involved in vesicle transport and protein trafficking, the NIS PM complexes were particularly enriched in proteins associated with the regulation of the actin cytoskeleton. Through a systematic validation of the detected interactions by co-immunoprecipitation and Western blot, followed by the biochemical and functional characterization of the contribution of each interactor to NIS PM residency and iodide uptake, we were able to identify a pathway by which the PM localization and function of NIS depends on its binding to SRC kinase, which leads to the recruitment and activation of the small GTPase RAC1. RAC1 signals through PAK1 and PIP5K to promote ARP2/3-mediated actin polymerization, and the recruitment and binding of the actin anchoring protein EZRIN to NIS, promoting its residency and function at the PM of normal and TC cells. Besides providing novel insights into the regulation of NIS localization and function at the PM of TC cells, our results open new venues for therapeutic intervention in TC, namely the possibility of modulating abnormal SRC signaling in refractory TC from a proliferative/invasive effect to the re-sensitization of these tumors to RAI therapy by inducing NIS retention at the PM.pt_PT
dc.description.sponsorshipThis work was funded by Bolsa Edward Limbert Merck/SPEDM -2021 (from Merck/SPEDM, Portugal) and Fundação para a Ciência e a Tecnologia (FCT), Portugal, through grants PTDC/BIAMOL/31787/2017 (to ALS and PM) and UID/MULTI/04046/2019 (to BioISI). MF was the recipient of FCT fellowship PD/BD/114388/2016, within the BioSYS PhD programme from BioISI.pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationCancers (Basel). 2021 Oct 30;13(21):5460. doi: 10.3390/cancers13215460pt_PT
dc.identifier.doi10.3390/cancers13215460pt_PT
dc.identifier.issn2072-6694
dc.identifier.urihttp://hdl.handle.net/10400.18/7926
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherMDPIpt_PT
dc.relationPTDC/BIAMOL/31787/2017pt_PT
dc.relationBiosystems & Integrative Sciences Institute
dc.relationTargeting Rac1-signaling to enhance iodide-related cancer therapy
dc.relation.publisherversionhttps://www.mdpi.com/2072-6694/13/21/5460pt_PT
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/pt_PT
dc.subjectThyroid Cancerpt_PT
dc.subjectNISpt_PT
dc.subjectRAC1 Signalingpt_PT
dc.subjectRAI-refractorypt_PT
dc.subjectPlasma Membrane Localizationpt_PT
dc.subjectGTPase Signalingpt_PT
dc.subjectTumorigenesispt_PT
dc.subjectVias de Transdução de Sinal e Patologias Associadaspt_PT
dc.titleAnalysis of NIS Plasma Membrane Interactors Discloses Key Regulation by a SRC/RAC1/PAK1/PIP5K/EZRIN Pathway with Potential Implications for Radioiodine Re-Sensitization Therapy in Thyroid Cancerpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.awardTitleBiosystems & Integrative Sciences Institute
oaire.awardTitleTargeting Rac1-signaling to enhance iodide-related cancer therapy
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UID%2FMulti%2F04046%2F2019/PT
oaire.awardURIinfo:eu-repo/grantAgreement/FCT//PD%2FBD%2F114388%2F2016/PT
oaire.citation.issue21pt_PT
oaire.citation.startPage5460pt_PT
oaire.citation.titleCancerspt_PT
oaire.citation.volume13pt_PT
oaire.fundingStream6817 - DCRRNI ID
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
project.funder.nameFundação para a Ciência e a Tecnologia
rcaap.embargofctAcesso de acordo com política editorial da revista.pt_PT
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT
relation.isProjectOfPublication35168786-8dfc-4a00-9759-dab3669fe1ae
relation.isProjectOfPublication266a9c6d-196d-4e29-92fb-3b9df1f2cf64
relation.isProjectOfPublication.latestForDiscovery35168786-8dfc-4a00-9759-dab3669fe1ae

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