CAJ | 학술논문

目的：研究支原体感染对人类细胞株的核糖体新生肽链复合体（ RNC）中 rRNA组成的影响及其相关机制。方法：分别提取支原体阳性与阴性RNC-RNA进行琼脂糖凝胶电泳，分析RNC-rRNA组成改变情况；在支原体污染环境中培养人正常肺上皮（ HBE）细胞，比较培养前后RNC-rRNA的变化；对RNC-RNA条带异常细胞进行抗支原体治疗，比较治疗前后RNC-rRNA的变化。利用免疫印迹分析支原体污染对丝裂原活化蛋白激酶（ mito-gen-activated protein kinase， MAPK）途径的影响。结果：不同组织来源的多株细胞中，支原体阴性和阳性细胞RNC-RNA的琼脂糖电泳显示细胞中RNC-rRNA组成发生异常改变，主要体现为28S和18S真核rRNA的降低，以及16S和未知原核rRNA的增加。在污染环境中培养HBE细胞，其RNC-rRNA组成可由支原体阴性转变为阳性谱型。以环丙沙星抗支原体治疗可逆转上述RNC-rRNA谱型的改变。总蛋白和磷酸化蛋白的免疫印记结果表明，支原体污染显著抑制A549细胞的p38 MAPK途径的活化，而对该细胞的ERK1／2途径无显著改变。结论：支原体感染可通过抑制p38 MAPK活化严重改变人类细胞株中RNC-rRNA的组成，从而影响宿主细胞的翻译行为。
목적：연구지원체감염대인류세포주적핵당체신생태련복합체（ RNC）중 rRNA조성적영향급기상관궤제。방법：분별제취지원체양성여음성RNC-RNA진행경지당응효전영，분석RNC-rRNA조성개변정황；재지원체오염배경중배양인정상폐상피（ HBE）세포，비교배양전후RNC-rRNA적변화；대RNC-RNA조대이상세포진행항지원체치료，비교치료전후RNC-rRNA적변화。이용면역인적분석지원체오염대사렬원활화단백격매（ mito-gen-activated protein kinase， MAPK）도경적영향。결과：불동조직래원적다주세포중，지원체음성화양성세포RNC-RNA적경지당전영현시세포중RNC-rRNA조성발생이상개변，주요체현위28S화18S진핵rRNA적강저，이급16S화미지원핵rRNA적증가。재오염배경중배양HBE세포，기RNC-rRNA조성가유지원체음성전변위양성보형。이배병사성항지원체치료가역전상술RNC-rRNA보형적개변。총단백화린산화단백적면역인기결과표명，지원체오염현저억제A549세포적p38 MAPK도경적활화，이대해세포적ERK1／2도경무현저개변。결론：지원체감염가통과억제p38 MAPK활화엄중개변인류세포주중RNC-rRNA적조성，종이영향숙주세포적번역행위。
AIM:To evaluate the effect of mycoplasma on the rRNA composition of mature ribosomes in human cell lines.METHODS:We isolated ribosome nascent-chain complex ( RNC) from multiple mycoplasma-positive-negative human cell lines.RNC-RNA was acquired from each cell line through RNA extraction, followed by agarose gel separation, fluorescent visualization and gray-scale value measurements on the rRNA bands.Western blot analysis was performed to in-vestigate the MAPK pathway alterations.RESULTS:In various human cell lines derived from different tissues, we found that as compared with mycoplasma-negative cells, mycoplasma-positive cells showed aberrant RNC-rRNA band patterns, featured by the decreases in 28S and 18S eukaryotic rRNAs and the increases in 16S and other unknown prokaryotic rRNAs.When cultured without ciprofloxacin, mycoplasma-negative HBE cells acquired mycoplasma contamination as ob-served with such characteristic abnormal rRNA bands.The ciprofloxacin treatment was able to recover the RNC-rRNA bands of the mycoplasma-contaminated cells to the normal pattern.The results of Western blot analysis on total and phos-phorylated proteins indicated that p38 pathway was significantly deactivated in mycoplasma-infected A549 cells, while ERK1/2 pathway was not significantly altered.CONCLUSION:Mycoplasma contamination severely alters the RNC-rRNA composition via p38 MAPK pathway, thus seriously impacting on the host cell translational behaviors.