Antisense transcripts play an important role in generating regulatory non-coding RNAs but whether these transcripts are also translated to generate functional peptides remains poorly understood. In this study, RNA sequencing and six-frame database generation were combined with mass spectrometry analysis of peptides isolated from polysomes to identify Nascent Pioneer Translation Products (Na-PTPs) originating from alternative reading frames of bi-directional transcripts. Two Na-PTP originating peptides derived from antisense strands stimulated CD8+ T cell proliferation when presented to peripheral blood mononuclear cells (PBMCs) from nine healthy donors. Importantly, an antigenic peptide derived from the reverse strand of two cDNA constructs was presented on MHC-I molecules and induced CD8+ T cell activation. The results demonstrate that three-frame translation of bi-directional transcripts generates antigenic peptide substrates for the immune system. This discovery holds significance for understanding the origin of self-discriminating peptide substrates for the major histocompatibility class I (MHC-I) pathway and for enhancing immune-based therapies against infected or transformed cells.

Department of Medical Biosciences Umeå University Umeå Sweden

Inserm UMRS1131 Institut de Génétique Moléculaire Université Paris Cité Hôpital St Louis Paris France

Laboratory of Growth Regulators Institute of Experimental Botany The Czech Academy of Sciences Olomouc Czechia

RECAMO Masaryk Memorial Cancer Institute Brno Czechia

UMR 1015 Immunologie des tumeurs et immunothérapie contre le cancer B2M Gustave Roussy Université Paris Sud Villejuif France

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