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Ｏｂｊｅｃｔｉｖｅ　Ｔｏ ｉｎｖｅｓｔｉｇａｔｅ ｔｈｅ ｅｆｆｅｃｔ ｏｆ Δ４０ｐ５３， ａｎ ａｌｔｅｒｎａｔｉｖｅ ｓｐｌｉｃｅｄ ｉｓｏｆｏｒｍ ｏｆ ｐ５３ ｌａｃｋｉｎｇ ｔｈｅ Ｎ⁃ｔｅｒ ｍｉｎｕｓ， ｏｎ ｔｈｅ ｐｒｏ⁃ａｐｏｐｔｏｔｉｃ ｆｕｎｃｔｉｏｎ ｏｆ ｐ５３． Ｍｅｔｈｏｄｓ　Ｔｈｅ ｗｉｌｄ⁃ｔｙｐｅ ｐ５３ ｗａｓ ｅｃｔｏｐｉｃａｌｌｙ ｅｘｐｒｅｓｓｅｄ ｉｎ ＨＣＴ１１６⁃ｐ５３－／ －（ｅｎｄｏｇｅｎｏｕｓ Δ４０ｐ５３ ｅｘｐｒｅｓｓｉｏｎ）， ＨＣＴ１１６⁃ｐ５３＋／ ＋（ｗｉｌｄ⁃ｔｙｐｅ ｐ５３） ａｎｄ Ｈ１２９９ （ｐ５３⁃ｎｕｌｌ） ｃｅｌｌｓ ｂｙ ａｄｅｎｏｖｉｒａｌ ｄｅｌｉｖｅｒｙ， ｗｈｉｌｅ Δ４０ｐ５３ ｐｌａｓｍｉｄ ｗｅｒｅ ｔｒａｎｓｆｅｃｔｅｄ ｉｎｔｏ ｔｈｅｓｅ ｃｅｌｌｓ ｔｏ ｏｖｅｒｅｘｐｒｅｓｓ Δ４０ｐ５３． Ｔｈｅ ｌｅｖｅｌｓ ｏｆ Δ４０ｐ５３ ａｎｄ ｐ５３ ｍＲＮＡ ｗｅｒｅ ｄｅｔｅｃｔｅｄ ｂｙ ｒｅｖｅｒｓｅ ｔｒａｎｓｃｒｉｐｔｉｏｎ⁃ｐｏｌｙｍｅｒａｓｅ ｃｈａｉｎ ｒｅａｃｔｉｏｎ （ＲＴ⁃ＰＣＲ） ａｎｄ ｑｕａｎｔｉｔａｔｉｖｅ ＰＣＲ． Ｔｈｅ ｅｘｐｒｅｓｓｉｏｎ ｏｆ ｒｅｌａｔｅｄ ｐｒｏｔｅｉｎｓ ｗａｓ ｄｅｔｅｒ ｍｉｎｅｄ ｂｙ Ｗｅｓｔｅｒｎ ｂｌｏｔｔｉｎｇ． Ｔｈｅ ｉｎｔｅｒａｃｔｉｏｎ ｏｆ ｐ５３ ａｎｄ Δ４０ｐ５３ ｗａｓ ｏｂｓｅｒｖｅｄ ｂｙ ｃｏ⁃ｉｍｍｕｎｏｐｒｅｃｉｐｉｔａｔｉｏｎ ａｓｓａｙ． Ｃａｌｃｅｉｎ⁃ＡＭ／ ｐｒｏｐｉｄｉｕｍ ｉｏｄｉｄｅ （ＰＩ） ｓｔａｉｎｉｎｇ ａｎｄ ｆｌｏｗ ｃｙｔｏｍｅｔｒｙ ｗｅｒｅ ｕｓｅｄ ｔｏ ｄｅｔｅｃｔ ｔｈｅ ａｐｏｐｔｏｔｉｃ ｒａｔｅ ｏｆ ｔｅｓｔｅｄ ｃｅｌｌｓ ｉｎ ｅａｃｈ ｇｒｏｕｐ． Ｒｅｓｕｌｔｓ　ＨＣＴ１１６⁃ｐ５３－／ － ｃｅｌｌｓ ｅｘｐｒｅｓｓｅｄ ｅｎｄｏｇｅｎｏｕｓ Δ４０ｐ５３ ｉｓｏｆｏｒｍ． Ｎｅｉｔｈｅｒ ｔｒａｎｓｃｒｉｐｔｉｏｎ ｎｏｒ ｐｒｏｔｅｉｎ ｅｘｐｒｅｓｓｉｏｎ ｏｆ ｗｉｌｄ⁃ｔｙｐｅ ｐ５３ ｗａｓ ｉｎｔｅｒｆｅｒｅｄ ｂｙ ｔｈｅ ｉｎｃｒｅａｓｅｄ ｅｘｐｒｅｓｓｉｏｎ ｏｆ Δ４０ｐ５３． Ｆｕｌｌ ｌｅｎｇｔｈ ｐ５３ ａｎｄ Δ４０ｐ５３ ｃｏｕｌｄ ｂｉｎｄ ｔｏ ｅａｃｈ ｏｔｈｅｒ． Ｃａｌｃｅｉｎ⁃ＡＭ／ ＰＩ ｓｔａｉｎｉｎｇ ｓｈｏｗｅｄ ｔｈａｔ ｔｈｅ ａｐｏｐｔｏｔｉｃ ｒａｔｅｓ ｏｆ Ｈ１２９９⁃Ｃｏｎｔｒｏｌ， ＨＣＴ１１６⁃ｐ５３－／ － ⁃Ｃｏｎｔｒｏｌ， Ｈ１２９９＋ｐ５３， ＨＣＴ１１６⁃ｐ５３－／ － ＋ｐ５３， Ｈ１２９９＋ｏｘａｌｉｐｌａｔｉｎ （Ｏｘａ）， ＨＣＴ１１６⁃ｐ５３－／ － ＋Ｏｘａ， Ｈ１２９９＋ｐ５３＋Ｏｘａ ａｎｄ ＨＣＴ１１６⁃ｐ５３－／ － ＋ｐ５３＋Ｏｘａ ｇｒｏｕｐｓ ｗｅｒｅ （２．５０±０．４７）％， （２．４０±０．３２）％， （５．２０±０．５８）％，（４．１０±０．１８）％， （２２．４０±１．７３）％， （１９．３０±１．１１）％， （２９．９０±１．１５）％ ａｎｄ （３９．３０±２．２６）％， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｉｔ ｗａｓ ｓｔａｔｉｓｔｉｃａｌｌｙ ｓｉｇｎｉｆｉｃａｎｔ ｂｅｔｗｅｅｎ Ｈ１２９９＋ｐ５３＋Ｏｘａ ａｎｄ ＨＣＴ１１６⁃ｐ５３－／ － ＋ ｐ５３＋Ｏｘａ ｇｒｏｕｐｓ （ｔ ＝ ３．７２１， Ｐ ＝ ０．０２０５）． Ｍｏｒｅｏｖｅｒ， ｔｈｅ ａｐｏｐｔｏｔｉｃ ｒａｔｅｓ ｏｆ Ｈ１２９９⁃Ｃｏｎｔｒｏｌ， Ｈ１２９９＋Δ４０ｐ５３， Ｈ１２９９＋ｐ５３， Ｈ１２９９＋ｐ５３＋Δ４０ｐ５３， Ｈ１２９９＋Ｏｘａ， Ｈ１２９９＋Δ４０ｐ５３＋Ｏｘａ， Ｈ１２９９＋ｐ５３＋Ｏｘａ ａｎｄ Ｈ１２９９＋ｐ５３＋Δ４０ｐ５３＋Ｏｘａ ｇｒｏｕｐｓ ｗｅｒｅ （２．６０±０．３５）％， （２．２０±０．１７）％， （４．８０±０．４９）％， （４．９０±１．１０）％， （２０．３０±１．１０）％， （１９．６０±１．４５）％， （２７．９０±１．３９）％， （３５．２０±１．４３）％， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｆｕｒｔｈｅｒｍｏｒｅ， ｆｌｏｗ ｃｙｔｏｍｅｔｒｙ ａｓｓａｙ ｓｈｏｗｅｄ ｔｈａｔ ｔｈｅ ａｐｏｐｔｏｔｉｃ ｒａｔｅｓ ｏｆ ａｂｏｖｅ ｃｅｌｌｓ ｗｅｒｅ （２．７０± ０．３２）％， （２．２０± ０．２４）％， （４．６０±０．４８）％， （３．９０±０．６７）％， （１９．３０±１．１１）％， （１７．７０±０．６６）％， （２８．３０±２．７６）％ ａｎｄ （３７．５０± １．５１）％， ｒｅｓｐｅｃｔｉｖｅｌｙ． Ｈ１２９９＋ｐ５３＋Δ４０ｐ５３＋Ｏｘａ ｃｅｌｌｓ ｓｈｏｗｅｄ ｈｉｇｈｅｒ ｃｅｌｌ ａｐｏｐｔｏｓｉｓ ｔｈａｎ Ｈ１２９９＋ｐ５３＋Ｏｘａ ｃｅｌｌｓ （ｔ ＝２．９３０， Ｐ ＝ ０．０４２）． Ｃｏｎｃｌｕｓｉｏｎ　Δ４０ｐ５３ ｉｓｏｆｏｒｍ ｃａｎ ｂｉｎｄ ｔｏ ｆｕｌｌ⁃ｌｅｎｇｔｈ ｐ５３， ａｎｄ ｅｎｈａｎｃｅ ｉｔｓ ｐｒｏ⁃ａｐｏｐｔｏｔｉｃ ｆｕｎｃｔｉｏｎ ｉｎ ｔｕｍｏｒ ｃｅｌｌｓ． [ABSTRACT FROM AUTHOR]

Objective To investigate the relationships of T→C mutation of 3264 site in tRNAleu region of mitochondrial DNA (mtDNA) with tumorigenesis and p53 mutation. Methods The whole DNA was extracted from carcinoma and adjacent tissues of 28 patients with liver carcinoma and 35 patients with colon cancer. Then tRNAleu region of mtDNA and p53 gene were amplified by PCR and sequenced. Results No T→C mutation was found in 3264 site, but 16 cases of mutations in the other sites were discovered in the carcinoma tissues. No T→C mutation was found in 3264 site, but 3 cases of mutations were discovered in the adjacent tissues .Seventeen cases of p53 gene mutation were found in the carcinoma tissues; 9 cases of p53 mutation were found in patients with colon carcinoma , including 6 cases of tRNAleu mutation and 3 cases of non-tRNAleu mutation (P<0.05); and 8 cases of p53 mutation were found in patients with liver cancer , including 5 cases of tRNAleu mutation and 3 cases of non-tRNAleu mutation (P<0.05). Conclusions The T→C mutation in 3264 site of tRNAleu region can′t be considered to have relationship with tumorigenesis. The mutations in other sites of tRNAleu region are due to p53 mutation, so the gene loses its protection, and thus the mutation occurs. [ABSTRACT FROM AUTHOR]