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The SLE variant Ala71Thr of BLK severely decreases protein abundance and binding to BANK1 through impairment of the SH3 domain function

Genes & Immunity volume 17, pages 128–138 (2016)Cite this article

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Abstract

The B-lymphocyte kinase (BLK) gene is associated genetically with several human autoimmune diseases including systemic lupus erythematosus. We recently described that the genetic risk is given by two haplotypes: one covering several strongly linked single-nucleotide polymorphisms within the promoter of the gene that correlated with low transcript levels, and a second haplotype that includes a rare nonsynonymous variant (Ala71Thr). Here we show that this variant, located within the BLK SH3 domain, is a major determinant of protein levels. In vitro analyses show that the 71Thr isoform is hyperphosphorylated and promotes kinase activation. As a consequence, BLK is ubiquitinated, its proteasomal degradation enhanced and the average life of the protein is reduced by half. Altogether, these findings suggest that an intrinsic autoregulatory mechanism previously unappreciated in BLK is disrupted by the 71Thr substitution. Because the SH3 domain is also involved in protein interactions, we sought for differences between the two isoforms in trafficking and binding to protein partners. We found that binding of the 71Thr variant to the adaptor protein BANK1 is severely reduced. Our study provides new insights on the intrinsic regulation of BLK activation and highlights the dominant role of its SH3 domain in BANK1 binding.

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Acknowledgements

We would like to thank Raquel Marrero Díaz, Maria José Luque, Manuel Martínez Bueno and Teresa Requena for technical assistance and José Antonio López Escámez for blood samples. Technical personnel of the Oklahoma Rheumatic Disease Research Cores Center (ORDRCC) are acknowledged for availability of some of the cell lines used in this study. Support to MEAR was provided through the following grants: Instituto de Salud Carlos III (CP10/00552 and PI12/02558) partly supported by FEDER funds, the Fundación Ramón Areces, the Swedish Research Council, OCAST, the Alliance for Lupus Research and the NIH (GM103456-10). Written informed consent was obtained from all subjects enrolled in the study. The study was reviewed and approved by the Ethical Committee of the Hospital Virgen de la Macarena, Sevilla, Spain and by the Institutional Review Board of the Oklahoma Medical Research Foundation.

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  1. M E Alarcón-Riquelme and C Castillejo-López: Authors share senior position.

Authors and Affiliations

  1. Area of Medical Genomics, Centro de Genómica e Investigación Oncológica (GENYO), Pfizer-Universidad de Granada-Junta de Andalucía, PTS, Granada, Spain

    A Díaz-Barreiro, M Bernal-Quirós, I Georg, C Marañón & M E Alarcón-Riquelme

  2. Institute for Environmental Medicine, Karolinska Institutet, Stockholm, Sweden

    M E Alarcón-Riquelme

  3. Department of Medical Sciences, Molecular Epidemiology, Uppsala University, Uppsala, Sweden

    C Castillejo-López

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  1. A Díaz-Barreiro
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Díaz-Barreiro, A., Bernal-Quirós, M., Georg, I. et al. The SLE variant Ala71Thr of BLK severely decreases protein abundance and binding to BANK1 through impairment of the SH3 domain function. Genes Immun 17, 128–138 (2016). https://doi.org/10.1038/gene.2016.1

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