Antigen-specificity measurements are the key to understanding T cell responses.

Antigen-specific T cells play a central role in the adaptive immune response and come in a wide range of phenotypes. T cell receptors (TCRs) mediate the antigen-specificities found in T cells. Importantly, high-throughput TCR sequencing provides a fingerprint which allows tracking of specific T cells and their clonal expansion in response to particular antigens. As a result, many studies have leveraged TCR sequencing in an attempt to elucidate the role of antigen-specific T cells in various contexts. Here, we discuss the published approaches to studying antigen-specific T cells and their specific TCR repertoire. Further, we discuss how these methods have been applied to study the TCR repertoire in various diseases in order to characterize the antigen-specific T cells involved in the immune control of disease.

Expression of specific HLA class II alleles is associated with an increased risk for active tuberculosis and a distinct gene expression profile.

Several HLA allelic variants have been associated with protection from or susceptibility to infectious and autoimmune diseases. Here, we examined whether specific HLA alleles would be associated with different Mycobacterium tuberculosis (Mtb) infection outcomes. The HLA alleles present at the -A, -B, -C, -DPA1, -DPB1, -DQA1, -DQB1, -DRB1, and -DRB3/4/5 loci were determined in a cohort of 636 individuals with known Mtb infection outcomes from South Africa and the United States. Among these individuals, 203 were QuantiFERON (QFT) negative, and 433 were QFT positive, indicating Mtb exposure. Of these, 99 QFT positive participants either had active tuberculosis (TB) upon enrollment or were diagnosed in the past. We found that DQA1*03:01, DPB1*04:02, and DRB4*01:01 were significantly more frequent in individuals with active TB (susceptibility alleles), as judged by Odds Ratios and associated p-values, while DPB1*105:01 was associated with protection from active TB. Peripheral blood mononuclear cells (PMBCs) from a subset of individuals were stimulated with Mtb antigens, revealing individuals who express any of the three susceptibility alleles were associated with lower magnitude of responses. Furthermore, we defined a gene signature associated with individuals expressing the susceptibility alleles that was characterized by lower expression of APC-related genes. In summary, we have identified specific HLA alleles associated with susceptibility to active TB and found that the expression of these alleles was associated with a decreased Mtb-specific T cell response and a specific gene expression signature. These results will help understand individual risk factors in progressing to active TB.

Rapid Analysis of ADP-Ribosylation Dynamics and Site-Specificity Using TLC-MALDI.

Poly(ADP-ribose) polymerases, PARPs, transfer ADP-ribose onto target proteins from nicotinamide adenine dinucleotide (NAD+). Current mass spectrometric analytical methods require proteolysis of target proteins, limiting the study of dynamic ADP-ribosylation on contiguous proteins. Herein, we present a matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) method that facilitates multisite analysis of ADP-ribosylation. We observe divergent ADP-ribosylation dynamics for the catalytic domains of PARPs 14 and 15, with PARP15 modifying more sites on itself (+3-4 ADP-ribose) than the closely related PARP14 protein (+1-2 ADP-ribose)─despite similar numbers of potential modification sites. We identify, for the first time, a minimal peptide fragment (18 amino-acids) that is preferentially modified by PARP14. Finally, we demonstrate through mutagenesis and chemical treatment with hydroxylamine that PARPs 14/15 prefer acidic residues. Our results highlight the utility of MALDI-TOF in the analysis of PARP target modifications and in elucidating the biochemical mechanism governing PARP target selection.