TRPC6 Channels Are Required for Proliferation, Migration and Invasion of Breast Cancer Cell Lines by Modulation of Orai1 and Orai3 Surface Exposure

Jardín, Isaac; Diez-Bello, Raquel; López, José J.; Redondo, Pedro C.; Salido, Ginés M.; Smani, Tarik; Rosado, Juan A.

doi:10.3390/cancers10090331

Cited by 73 publications

(102 citation statements)

References 49 publications

(67 reference statements)

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“…Consistent with previous studies [3,6], Western blot analysis of whole-cell lysates from the non-tumoral breast epithelial MCF10A cell line and the estrogen receptor positive (ER + ) and TNBC cell lines MCF7 and MDA-MB-231, respectively, with a specific anti-human Orai1 antibody revealed a low expression of Orai1 in MCF10A cells and a significantly higher expression of this protein in breast cancer cells (Figure 1a,b; p < 0.05; n = 6). The increased expression of Orai1 in the breast cancer cell lines is consistent with the high expression of this protein in cancerous tissue [22].…”

Section: Expression and Interaction Of Orai1α And Ac8 In Non-tumoralsupporting

confidence: 93%

“…Orai1 was reported to play a major role in Ca 2+ influx in the TNBC cell line MDA-MB-231. SOCE, a major mechanism for Ca 2+ entry in this cell type, was found to be strongly dependent on Orai1 [6], whose plasma membrane expression is modulated by TRPC6 channels [3]. Furthermore, Orai1 is…”

Section: Discussionmentioning

confidence: 99%

“…SOCE in MDA-MB-231 cells was reported to be entirely dependent on Orai1 function [3,6]. Hence, we explored the functional role of the interaction between Orai1 and AC8 in these cells.…”

Section: Role Of Ac8 In the Activation Of Store-operated Ca 2+ Entrymentioning

confidence: 99%

“…Ca 2+ influx via Orai1 channels was reported to play a relevant role in the development of a number of cancer hallmarks, including cell migration [3,[26][27][28]. Next, we assessed the relevance of AC8 in the ability of these cell lines to migrate.…”

Section: Ac8 Plays a Relevant Role In Mda-mb-231 Cell Migrationmentioning

confidence: 99%

“…Consequently, this type of cancer is resistant to hormonal therapies and chemicals that target the HER2 receptor [1]. Studies in TNBC cells revealed that Ca 2+ signaling is remodeled and plays a key functional role [2][3][4][5]. TNBC cells overexpress Orai1, which is responsible for the activation of store-operated Ca 2+ entry (SOCE) [6].…”

Section: Introductionmentioning

confidence: 99%

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Adenylyl Cyclase Type 8 Overexpression Impairs Phosphorylation-Dependent Orai1 Inactivation and Promotes Migration in MDA-MB-231 Breast Cancer Cells

Sánchez-Collado

López

Jardín

et al. 2019

Cancers

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Orai1 plays a major role in store-operated Ca 2+ entry (SOCE) in triple-negative breast cancer (TNBC) cells. This channel is inactivated via different mechanisms, including protein kinase C (PKC) and protein kinase A (PKA)-dependent phosphorylation at Ser-27 and Ser-30 or Ser-34, respectively, which shapes the Ca 2+ responses to agonists. The Ca 2+ calmodulin-activated adenylyl cyclase type 8 (AC8) was reported to interact directly with Orai1, thus mediating a dynamic interplay between the Ca 2+ -and cyclic adenosine monophosphate (cAMP)-dependent signaling pathways.Here, we show that the breast cancer cell lines MCF7 and MDA-MB-231 exhibit enhanced expression of Orai1 and AC8 as compared to the non-tumoral breast epithelial MCF10A cell line. In these cells, AC8 interacts with the Orai1α variant in a manner that is not regulated by Orai1 phosphorylation. AC8 knockdown in MDA-MB-231 cells, using two different small interfering RNAs (siRNAs), attenuates thapsigargin (TG)-induced Ca 2+ entry and also Ca 2+ influx mediated by co-expression of Orai1 and the Orai1-activating small fragment (OASF) of STIM1 (stromal interaction molecule-1). Conversely, AC8 overexpression enhances SOCE, as well as Ca 2+ entry, in cells co-expressing Orai1 and OASF. In MDA-MB-231 cells, we found that AC8 overexpression reduces the Orai1 phosphoserine content, thus suggesting that AC8 interferes with Orai1 serine phosphorylation, which takes place at residues located in the AC8-binding site. Consistent with this, the subset of Orai1 associated with AC8 in naïve MDA-MB-231 cells is not phosphorylated in serine residues in contrast to the AC8-independent Orai1 subset. AC8 expression knockdown attenuates migration of MCF7 and MDA-MB-231 cells, while this maneuver has no effect in the MCF10A cell line, which is likely attributed to the low expression of AC8 in these cells. We found that AC8 is required for FAK (focal adhesion kinase) phosphorylation in MDA-MB-231 cells, which might explain its role in cell migration. Finally, we found that AC8 is required for TNBC cell proliferation. These findings indicate that overexpression of AC8 in breast cancer MDA-MB-231 cells impairs the phosphorylation-dependent Orai1 inactivation, a mechanism that might support the enhanced ability of these cells to migrate.

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Section: Expression and Interaction Of Orai1α And Ac8 In Non-tumoralsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

“…SOCE in MDA-MB-231 cells was reported to be entirely dependent on Orai1 function [3,6]. Hence, we explored the functional role of the interaction between Orai1 and AC8 in these cells.…”

Section: Role Of Ac8 In the Activation Of Store-operated Ca 2+ Entrymentioning

confidence: 99%

Section: Ac8 Plays a Relevant Role In Mda-mb-231 Cell Migrationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Adenylyl Cyclase Type 8 Overexpression Impairs Phosphorylation-Dependent Orai1 Inactivation and Promotes Migration in MDA-MB-231 Breast Cancer Cells

Sánchez-Collado

López

Jardín

et al. 2019

Cancers

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Postbiotics of Lacticaseibacillus paracaseiCECT 9610 and Lactiplantibacillus plantarumCECT 9608 attenuates store‐operated calcium entry and FAK phosphorylation in colorectal cancer cells

Macias‐Diaz,

Lopez,

Bravo

et al. 2024

Molecular Oncology

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Store‐operated Ca2+ entry (SOCE) is a major mechanism for Ca2+ influx in colorectal cancer (CRC) cells. This mechanism, regulated by the filling state of the intracellular Ca2+ stores, is mediated by the endoplasmic reticulum Ca2+ sensors of the stromal interaction molecules (STIM) family [stromal interaction molecule 1 (STIM1) and STIM2] and the Ca2+‐release‐activated Ca2+ channels constituted by Orai family members, with predominance of calcium release‐activated calcium channel protein 1 (Orai1). CRC cells exhibit enhanced SOCE due to remodeling of the expression of the key SOCE molecular components. The enhanced SOCE supports a variety of cancer hallmarks. Here, we show that treatment of the colorectal adenocarcinoma cell lines HT‐29 and Caco‐2 with inanimate Lacticaseibacillus paracasei (CECT9610) and Lactiplantibacillus plantarum (CECT9608) attenuates SOCE, although no detectable effect is seen on SOCE in normal colon mucosa cells. The effect of Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics was mediated by downregulation of Orai1 and STIM1, while the expression levels of Orai3 and STIM2 remained unaltered. Treatment of HT‐29 and Caco‐2 cells with inanimate Lacticaseibacillus paracasei and Lactiplantibacillus plantarum impairs in vitro migration by a mechanism likely involving attenuation of focal adhesion kinase (FAK) tyrosine phosphorylation. Cell treatment with the Orai1 inhibitor synta‐66 attenuates SOCE and prevents any further effect of Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics. Together, our results indicate for the first time that Lacticaseibacillus paracasei and Lactiplantibacillus plantarum postbiotics selectively exert negative effects on Ca2+ influx through SOCE in colorectal adenocarcinoma cell lines, providing evidence for an attractive strategy against CRC.

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Functional differences in agonist‐induced plasma membrane expression of Orai1α and Orai1β

Jardín

Alvarado

Sánchez-Collado

et al. 2023

Journal Cellular Physiology

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Orai1 is the pore‐forming subunit of the store‐operated Ca2+ release‐activated Ca2+ (CRAC) channels involved in a variety of cellular functions. Two Orai1 variants have been identified, the long form, Orai1α, containing 301 amino acids, and the short form, Orai1β, which arises from alternative translation initiation from methionines 64 or 71, in Orai1α. Orai1 is mostly expressed in the plasma membrane, but a subset of Orai1 is located in intracellular compartments. Here we show that Ca2+ store depletion leads to trafficking and insertion of compartmentalized Orai1α in the plasma membrane via a mechanism that is independent on changes in cytosolic free‐Ca2+ concentration, as demonstrated by cell loading with the fast intracellular Ca2+ chelator dimethyl BAPTA in the absence of extracellular Ca2+. Interestingly, thapsigargin (TG) was found to be unable to induce translocation of Orai1β to the plasma membrane when expressed individually; by contrast, when Orai1β is co‐expressed with Orai1α, cell treatment with TG induced rapid trafficking and insertion of compartmentalized Orai1β in the plasma membrane. Translocation of Orai1 forms to the plasma membrane was found to require the integrity of the actin cytoskeleton. Finally, expression of a dominant negative mutant of the small GTPase ARF6, and ARF6‐T27N, abolished the translocation of compartmentalized Orai1 variants to the plasma membrane upon store depletion. These findings provide new insights into the mechanism that regulate the plasma membrane abundance of Orai1 variants after Ca2+ store depletion.

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TRPC6 Channels Are Required for Proliferation, Migration and Invasion of Breast Cancer Cell Lines by Modulation of Orai1 and Orai3 Surface Exposure

Cited by 73 publications

References 49 publications

Adenylyl Cyclase Type 8 Overexpression Impairs Phosphorylation-Dependent Orai1 Inactivation and Promotes Migration in MDA-MB-231 Breast Cancer Cells

Adenylyl Cyclase Type 8 Overexpression Impairs Phosphorylation-Dependent Orai1 Inactivation and Promotes Migration in MDA-MB-231 Breast Cancer Cells

Postbiotics of Lacticaseibacillus paracaseiCECT 9610 and Lactiplantibacillus plantarumCECT 9608 attenuates store‐operated calcium entry and FAK phosphorylation in colorectal cancer cells

Functional differences in agonist‐induced plasma membrane expression of Orai1α and Orai1β

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