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In Vitro Ubiquitination and Deubiquitination Assays of Nucleosomal Histones
Published on: July 25, 2019
Ubiquitinación en los residuos de no lisina por una ligasa viral E3 ubiquitina ligasa
1Department of Molecular and Cell Biology, 142 Life Sciences Addition Room 3200, Berkeley, CA 94720, USA.
Science (New York, N.Y.)
|July 5, 2005
Resumen
La ligasa MIR1 E3 del herpesvirus asociado al sarcoma de Kaposi se dirige a MHC I para la degradación, incluso sin residuos de lisina. Esta nueva vía de ubiquitinación involucra residuos de cisteína, ampliando nuestra comprensión de la regulación de las proteínas.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- Virología Virología.
- Biología celular Biología celular.
Sus antecedentes:
- La ubiquitinación es un proceso celular clave que regula la función de las proteínas, principalmente mediada por las ligasas de ubiquitina E3.
- Las ligasas de ubiquitina E3 determinan la especificidad del sustrato y catalizan la formación de enlaces isopéptidos entre la ubiquitina y los residuos de lisina del sustrato o el N-terminal.
- El herpesvirus asociado al sarcoma de Kaposi codifica las ligasas MIR1 y MIR2 E3 involucradas en la ubiquitinación e internalización del complejo mayor de histocompatibilidad clase I (MHC I).
Objetivo del estudio:
- Investigar el mecanismo por el cual las ligasas MIR1 y MIR2 E3 regulan la ubiquitinación y la regulación a la baja del MHC I.
- Determinar si MIR1 puede mediar la ubiquitinación y degradación de MHC I en ausencia de residuos de lisina accesibles.
Principales métodos:
- Utilizó moléculas MHC I mutantes que carecen de residuos de lisina intracitoplasmática.
- Estado de ubiquitinación evaluado de MHC I en presencia de MIR1 y MIR2.
- Se analizó la sensibilidad de la asociación ubiquitina-MHC I al beta2-mercaptoetanol.
- Investigó el papel de los residuos de cisteína en el proceso de ubiquitinación.
- Empleó moléculas MHC I diseñadas con dominios intracitoplasmáticos artificiales que contienen residuos de cisteína.
Principales resultados:
- MIR1, pero no MIR2, regulación descendente inducida de las moléculas MHC I que carecen de residuos de lisina intracitoplasmática.
- La ubiquitinación de estas moléculas MHC I deficientes en lisina por MIR1 fue sensible al beta2-mercaptoetanol, lo que indica un enlace no lisino.
- Esta nueva ubiquitinación requería un residuo de cisteína en la cola intracitoplasmática del MHC I.
- Una molécula de MHC I diseñada con una cisteína en una cola artificial fue endocitada y degradada por MIR1.
Conclusiones:
- MIR1 media una forma única de ubiquitinación dirigida a las proteínas que carecen de lisinas o N-terminales accesibles.
- Esta vía de ubiquitinación dependiente de la cisteína contribuye a la regulación a la baja del MHC I.
- Los hallazgos amplían los mecanismos conocidos de ubiquitinación de proteínas y la regulación por las ligasas virales E3.
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