Estudio de la regulación y función del canal orai1 en la homeostasis del calcio intracelular
- Autores: Joel Nieto Felipe
- Directores de la Tesis: Juan A. Rosado (dir. tes.), Isaac Jardín Polo (codir. tes.)
- Lectura: En la Universidad de Extremadura ( España ) en 2025
- Idioma: español
- Tribunal Calificador de la Tesis: Tarik Smani Hajami (presid.), José Javier López Barba (secret.), Rainer Schindl (voc.)
- Programa de doctorado: Programa de Doctorado en Biomarcadores de Salud y Estados Patológicos por la Universidad de Extremadura
- Materias:
- Enlaces
- Resumen
- español
La entrada capacitativa de Ca2+ (ECC) descrita en el año 1986 por James Putney Jr. (1) constituye el principal mecanismo de movilización del ion Ca2+ en las células no excitables, aunque también se ha descrito en células excitables. Este mecanismo está mediado por los canales de Ca2+ pertenecientes a la familia Orai, así como por las proteínas de la familia STIM que actúan como sensores de la concentración de Ca2+ en el retículo endoplásmico (RE). Cabe destacar que en el año 2012, se describió la existencia de dos variantes del canal, Orai1 alpha y Orai1 beta (2,3). Dada la importancia que tiene el ion Ca2+ en la regulación de procesos fisiológicos y celulares, como la transcripción génica así como en procesos patológicos como el cáncer, la ECC es un mecanismo que ha de encontrarse altamente regulado. Es por ello, que desde su descripción, varios han sido los estudios relacionados con el mecanismo de ECC que han descrito la participación de proteínas auxiliares implicadas en la regulación de la ECC (4).
Es por esto por lo que mediante el uso de microscopía de fluorescencia y confocal, ensayos de análisis de expresión génica por medio de emisión de bioluminiscencia, análisis de la capacidad migratoria, supervivencia y proliferación celular y análisis de la expresión proteica por diferentes técnicas hemos profundizado en el mecanismo de ECC, así como la participación de algunas de las proteínas relacionadas en la respuesta inmune o el desarrollo tumoral, entre otros mecanismo. La variante Orai1 alpha junto con STIM2 presenta una menor efectividad a la hora de mediar ECC en conjunto con STIM2 que Orai1 beta. Además, STIM1 y STIM2, son capaces de regular la expresión de las variantes de Orai1 por medio de la vía de degradación proteica endolisosomal. Al mismo tiempo, las proteínas reguladoras de ECC, SARAF y EFHB, son muy importantes para el correcto desarrollo de procesos tumorales tales como la supervivencia celular, capacidad de migración, proliferación y progresión del ciclo celular en células tumorales mamarias del subtipo luminal A y triple negativo. Finalmente, se ha descrito que la variantes Orai1 alpha es capaz de mediar la activación del factor de transcripción proinflamatorio NF-B de una forma dependiente de la movilización de Ca2+ y que requiere de la interacción del canal con la PKC beta 2.
Estos resultados ponen de manifiesto la importancia existente en la regulación de la ECC, dado que una desregulación de la misma puede desencadenar el desarrollo de situaciones patológicas como el cáncer o incluso una disminución de la capacidad de respuesta inflamatoria cuando esta sea necesaria.
1. Putney JW. A model for receptor-regulated calcium entry. Cell Calcium [Internet]. 1986 [cited 2023 Dec 4];7(1):1-12. Available from: https://pubmed.ncbi.nlm.nih.gov/2420465/ 2. Desai PN, Zhang X, Wu S, Janoshazi A, Bolimuntha S, Putney JW, et al. Multiple types of calcium channels arising from alternative translation initiation of the Orai1 message. Sci Signal [Internet]. 2015 Jul 28 [cited 2023 Dec 4];8(387). Available from: https://pubmed.ncbi.nlm.nih.gov/26221052/ 3. Fukushima M, Tomita T, Janoshazi A, Putney JW. Alternative translation initiation gives rise to two isoforms of Orai1 with distinct plasma membrane mobilities. J Cell Sci [Internet]. 2012 [cited 2023 Dec 4];125(Pt 18):4354-61. Available from: https://pubmed.ncbi.nlm.nih.gov/22641696/ 4. Lopez JJ, Jardin I, Albarrán L, Sanchez-Collado J, Cantonero C, Salido GM, et al. Molecular Basis and Regulation of Store-Operated Calcium Entry. Adv Exp Med Biol [Internet]. 2020 [cited 2024 Dec 18];1131:445-69. Available from: https://pubmed.ncbi.nlm.nih.gov/31646520/
- English
The Ca2+ ion is a widely distributed element in organisms whose main function is to act as a second messenger responsible for regulating a plethora of cellular functions and physiological processes. For this reason, cells must maintain a correct balance of ion concentrations both intracellularly and extracellularly, as an imbalance in either can have severe effects on all Ca2+-regulated functions as a second messenger. Many of these cellular functions require an increase in [Ca2+]c. At this point, there is a wide variety of channels that allow the entry of the ion from the extracellular environment, but we also find intracellular Ca2+ stores, mainly the ER, which have a series of receptors along their membrane, and the binding of their agonist will trigger their activation and thus the release of the ion into the cytosol. However, once cells have completed the specific function for which the increase in [Ca2+]c occurred, it is necessary to return these concentrations to resting conditions in order to avoid excessive and prolonged cytosolic Ca2+ overload, as this will be detrimental to the cells and the organism itself. This aspect necessitates a series of mechanisms capable of expelling Ca2+ back into the extracellular environment or, failing that, refilling previously emptied intracellular stores. These mechanisms include a series of channels, pumps, and exchangers that maintain this gradient between the different cellular compartments. The store-operated Ca2+ entry, described by James Putney Jr. in 1986, constitutes the main pathway for calcium mobilization in non-excitable cells, although it has also been described in excitable cells. In this case, the signaling cascade triggered by the binding of IP3 to its receptor in the ER promotes the emptying of the stores and thereby activates a calcium current that allows the ion to enter the cellular cytoplasm. At this point, the STIM family proteins act as luminal Ca2+ sensors that, depending on the filling state of the stores, will control the activation and opening of the capacitative channels through which the ion enters. These capacitative channels are formed by hexameric combinations of Orai family proteins, which are responsible for mediating highly selective currents for calcium known as ICRAC currents, although alternative currents or ISOC currents mediated by TRPC channels have been described but require some degree of activation of Orai1 channels. The importance of SOCE lies in its role as one of the main pathways for Ca2+ mobilization, which is crucial for regulating multiple physiological mechanisms such as gene transcription, immune response, platelet aggregation, etc. However, it also plays a significant role in pathological processes where dysregulation at any key point of the process, whether it's activation, interaction, inactivation, etc., of SOCE, can disrupt the proper functioning of these cellular functions or physiological processes, leading to various pathological conditions such as Stormorken syndrome or severe combined immunodeficiency caused by mutations in STIM and Orai proteins, respectively. Additionally, an increase in the expression of both proteins has been described in samples from cancer, among other diseases. This situation necessitates precise regulation of the SOCE mechanism at almost every level, with the main regulatory mechanism being Ca2+-dependent inactivation (CDI), involving proteins such as adenylate cyclase 8 (AC8) or calmodulin (CaM).Furthermore, in recent years, more proteins that could be involved in CDI have been identified. One of these is SARAF, described as a negative regulator, while our research group identified the EFHB protein, which appears to function as a positive regulator. The role of STIM2 in SOCE regulation is still not clear, making SARAF and EFHB together a good focus of study to establish the mechanical basis of SOCE andthus delve into aspects such as the role of Orai1α and Orai1β variants in regulating gene transcription associated with the inflammatory response. All of this has led to the present doctoral thesis being titled: "Study of the regulation and function of the Orai1 channel in intracellular calcium homeostasis", which will be submitted for evaluation by the tribunal for the obtaining of the doctorate degree from the University of Extremadura.
- español
