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Here, we combined antigen- specific T cell receptor (TCR) identification, longitudinal TCR sequencing, and single-cell multi- omics to characterize Env- and Gag-specific memory precursor T cells elicited by the HIV Vaccine Trials Network (HVTN) 505 DNA prime\u2013recombinant adenovirus serotype 5 (rAd5) boost (DNA\/rAd5) vaccine regimen. We developed a generalizable high-throughput approach to identify HIV-1 Env- and Gag-specific TCRs and found distinct patterns of clonal expansion, persistence, and contribution to memory between Env- and Gag-specific CD8\u207a T cell responses. The DNA prime and rAd5 boost differentially shaped these repertoires, with rAd5-induced clones contributing proportionally more to the Gag-specific than to the Env-specific memory precursor compartment. Single-cell immune profiling further revealed distinct memory precursor states, with Env-specific responses enriched for\n                  <jats:italic>\n                    GZMB\n                    <jats:sup>+<\/jats:sup>\n                    PRF1\n                    <jats:sup>+<\/jats:sup>\n                  <\/jats:italic>\n                  cytotoxic effector-memory (EM) CD8\u207a T cells and Gag-specific responses containing a larger cycling\/proliferative population. Together, these findings demonstrate that heterologous DNA\/rAd5 vaccination generates antigen-specific CD8\u207a T cell memory with distinct clonal trajectories and cellular programs, providing new insights into how vaccine platform and antigen-specificity shape the durability and functional properties of HIV-1 specific cellular immunity.\n                <\/jats:p>","DOI":"10.64898\/2026.09.23.752815","type":"posted-content","created":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T18:35:22Z","timestamp":1790706922000},"source":"Crossref","is-referenced-by-count":0,"title":["HIV-1 prime-boost vaccination shapes distinct clonal trajectories and memory precursor states of Env- and Gag-specific T cells"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7699-8837","authenticated-orcid":false,"given":"Guoyue","family":"Xu","sequence":"first","affiliation":[{"name":"Department of Global Health, University of Washington, Seattle, USA"},{"name":"Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3661-2071","authenticated-orcid":false,"given":"Joshua X.","family":"Yee","sequence":"additional","affiliation":[{"name":"Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1045-735X","authenticated-orcid":false,"given":"Saskia M.","family":"Ilcisin","sequence":"additional","affiliation":[{"name":"Allen Institute for Immunology, Seattle, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-8265-8169","authenticated-orcid":false,"given":"Saransh N.","family":"Kaul","sequence":"additional","affiliation":[{"name":"Allen Institute for Immunology, Seattle, 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A.","family":"Minervina","sequence":"additional","affiliation":[{"name":"Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5946-9733","authenticated-orcid":false,"given":"Shelly T.","family":"Karuna","sequence":"additional","affiliation":[{"name":"Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7112-3118","authenticated-orcid":false,"given":"Magdalena E.","family":"Sobieszczyk","sequence":"additional","affiliation":[{"name":"Aaron Diamond AIDS Research Center, Columbia University Vagelos College of Physicians and Surgeons, New York, USA"},{"name":"Department of Internal Medicine, Columbia University Vagelos College of Physicians and Surgeons, New York, 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