Major histocompatibility complex (MHC) class I polymorphisms influence outcomes in a number of infectious diseases, cancers and inflammatory diseases. In human immunodeficiency virus (HIV) infections, among all genetic factors known to influence progression to acquired immunodeficiency syndrome (AIDS), the strongest associations link to human MHC class I genes. MHC class I molecules bind to peptide antigens and present these antigens to CD8 T cells. MHC class I molecules are also ligands for inhibitory receptors of NK cells Three sets of genes encode human MHC class I molecules which are the human leukocyte antigens (HLA) A, B and C. These genes are highly polymorphic, with the HLA-B genes being the most variable. It is generally assumed that HLA-disease associations link to the peptide binding characteristics of individual HLA class I molecules. However, it remains largely unknown whether and how differences in assembly characteristics or stabilities of HLA class I molecules influence immunological outcomes. A set of objectives of this application is to examine such influences. A central hypothesis is that the observed assembly and stability differences between HLA-B allotypes influence their cell surface expression, and their abilities to mediate CD8 T cell and NK cell responses.
In Aim 1, we show that dependence on the assembly factor tapasin is quite variable among HLA-B molecules. Tapasin-dependent assembly is highly prevalent within the HLA-Bw4 serotype, whereas many HLA-B molecules of HLA-Bw6 serotype are tapasin-independent for their assembly. HLA-Bw4 molecules but not HLA-Bw6 molecules are ligands for inhibitory receptors of natural killer (NK) cells, which are responsive to cell surface expression densities of MHC class I molecules. To further examine whether the HLA-Bw4/HLA- Bw6 segregation in tapasin dependence reflects altered cell surface expression of different allotypes, quantitative flow cytometry will be used to compare cell surface expression densities of HLA-B molecules in primary human CD4 T cells under basal conditions, and following infections with HIV-1.
In Aim 2, we show varying conformational stabilities of soluble peptide-deficient HLA-B molecules during their refolding, and variable levels of expression of HLA-B molecules on cells deficient in the transporter associated with antigen processing (TAP). Based on these findings, we will examine variable recognition of HLA-B molecules by endoplasmic reticulum (ER) quality control factors, and differing requirements for other assembly factors. The molecular basis for stability differences of the empty HLA-B proteins will be elucidated.
In Aim 3, we will examine whether the observed HLA-B assembly and stability differences influence the breadth and stability of peptide selection by HLA-B molecules. These analyses will be undertaken by comparing different HLA-B- restricted CD8 T cell responses against peptide antigens spanning the HIV proteome. Taken together, these studies are significant towards defining variations in immune functions of different HLA-B molecules, which are relevant towards vaccine design and infectious disease outcomes.

Public Health Relevance

The studies described in this proposal are significant towards understanding how variations in human leukocyte antigens (HLA)-B influence their structure, cell surface expression and immune responses. These studies will ultimately be important towards vaccine design and immunotherapy that is individualized and based on the HLA genotype.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Cellular and Molecular Immunology - A Study Section (CMIA)
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Gondre-Lewis, Timothy A
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University of Michigan Ann Arbor
Schools of Medicine
Ann Arbor
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Geng, Jie; Zaitouna, Anita J; Raghavan, Malini (2018) Selected HLA-B allotypes are resistant to inhibition or deficiency of the transporter associated with antigen processing (TAP). PLoS Pathog 14:e1007171
Yarzabek, Brogan; Zaitouna, Anita J; Olson, Eli et al. (2018) Variations in HLA-B cell surface expression, half-life and extracellular antigen receptivity. Elife 7:
Geng, Jie; Altman, John D; Krishnakumar, Sujatha et al. (2018) Empty conformers of HLA-B preferentially bind CD8 and regulate CD8+ T cell function. Elife 7:
Wijeyesakere, Sanjeeva Joseph; Bedi, Sukhmani Kaur; Huynh, David et al. (2016) The C-Terminal Acidic Region of Calreticulin Mediates Phosphatidylserine Binding and Apoptotic Cell Phagocytosis. J Immunol 196:3896-3909
Wijeyesakere, Sanjeeva Joseph; Gagnon, Jessica K; Arora, Karunesh et al. (2015) Regulation of calreticulin-major histocompatibility complex (MHC) class I interactions by ATP. Proc Natl Acad Sci U S A 112:E5608-17
Geng, Jie; Pogozheva, Irina D; Mosberg, Henry I et al. (2015) Use of Functional Polymorphisms To Elucidate the Peptide Binding Site of TAP Complexes. J Immunol 195:3436-48
Deffit, Sarah N; Blum, Janice S (2015) A central role for HSC70 in regulating antigen trafficking and MHC class II presentation. Mol Immunol 68:85-8
Raghavan, Malini; Geng, Jie (2015) HLA-B polymorphisms and intracellular assembly modes. Mol Immunol 68:89-93
Wijeyesakere, Sanjeeva J; Rizvi, Syed M; Raghavan, Malini (2013) Glycan-dependent and -independent interactions contribute to cellular substrate recruitment by calreticulin. J Biol Chem 288:35104-16
Raghavan, Malini; Wijeyesakere, Sanjeeva J; Peters, Larry Robert et al. (2013) Calreticulin in the immune system: ins and outs. Trends Immunol 34:13-21

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