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The ATL-derived cell line MT-1 is used to study HTLV-1 biology and leukemogenesis. However, the proviral integration sites and internal structures of HTLV-1 are not comprehensively characterized in MT-1 cells. We analyzed two independently maintained MT-1 cell lines (MT-1\u00a0J and MT-1\u00a0M) using long PCR, quantitative PCR-based proviral load analysis, inverse long PCR, inverse PCR, integration site-specific PCR, direct sequencing, and Rapid Amplification of Integration Site without Interference by Genomic DNA contamination. Long PCR and proviral load analyses demonstrated that MT-1 cells harbor multiple full-length and defective HTLV-1 proviruses. MT-1\u00a0J cells exhibited reduced proviral load in the\n                    <jats:italic>pX<\/jats:italic>\n                    region, suggesting the presence of\n                    <jats:italic>pX<\/jats:italic>\n                    -lacking proviruses. Inverse long PCR and inverse PCR revealed at least five proviral integration sites in MT-1\u00a0J cells; MT-1\u00a0M cells contained three. Site-specific PCR confirmed the differential preservation of integration sites. Sequence analysis revealed two full-length proviruses and three type I defective proviruses with distinct internal deletions, including an unreported provirus with a large deletion encompassing\n                    <jats:italic>pX<\/jats:italic>\n                    . Rapid Amplification of Integration Site without Interference by Genomic DNA contamination identified major proviral clones shared between MT-1\u00a0J and MT-1\u00a0M cells, while revealing differences in clone frequencies and minor integration sites. The MT-1 cell line is a polyclonal population containing multiple full-length and defective HTLV-1 proviruses. Its proviral composition can change during long-term in vitro passaging. This heterogeneity should be considered when interpreting results obtained using MT-1 cells in HTLV-1 and ATL research.\n                  <\/jats:p>","DOI":"10.1007\/s13577-026-01385-1","type":"journal-article","created":{"date-parts":[[2026,4,30]],"date-time":"2026-04-30T18:13:07Z","timestamp":1777572787000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Heterogeneity of HTLV-1 proviral integration sites and internal structures in the ATL cell line MT-1"],"prefix":"10.1007","volume":"39","author":[{"given":"Misaki","family":"Izaki","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuki","family":"Hashikura","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1116-5818","authenticated-orcid":false,"given":"Kunihiko","family":"Umekita","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kazumi","family":"Umeki","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Taiga","family":"Miyazaki","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2026,4,30]]},"reference":[{"key":"1385_CR1","doi-asserted-by":"publisher","first-page":"268","DOI":"10.1038\/294268a0","volume":"294","author":"BJ Poiesz","year":"1981","unstructured":"Poiesz BJ, Ruscetti FW, Reitz MS, Kalyanaraman VS, Gallo RC. 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