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Overexpressing Long Noncoding RNAs Using Gene-activating CRISPR
Published on: March 1, 2019
Next-generation sequencing reveals differentially expressed small noncoding RNAs in uterine leiomyoma
Tsai-Der Chuang1, Yeming Xie2, Wei Yan2
1Department of Obstetrics and Gynecology, Harbor-UCLA Medical Center and LA-Biomed Research Institute, Torrance, California.
Objective:
To determine the expression profile of small noncoding RNAs (sncRNAs) in leiomyoma, which has not been investigated to date.
Design:
Laboratory-based investigation.
Setting:
Academic center.
Patient(S):
Women undergoing hysterectomy for benign indications.
Intervention(S):
Next-generation sequencing and screening of an sncRNA database with confirmatory analysis by quantitative reverse-transcription polymerase chain reaction (qRT-PCR).
Main Outcome Measure(S):
Expression profile of sncRNAs in leiomyoma and matched myometrium.
Result(S):
Screening our previously determined RNA sequencing data with the sncRNA database resulted in identification of 15 small nuclear (sn) RNAs, 284 small nucleolar (sno) RNAs, 98 Piwi-interacting (pi) RNAs, 152 transfer (t) RNAs, and 45 ribosomal (r) RNAs, of which 15 snoRNAs, 24 piRNAs, 7 tRNAs, and 6 rRNAs were differentially expressed at a 1.5-fold change cutoff in leiomyoma compared with myometrium. We selected 5 snoRNAs, 4 piRNAs, 1 tRNA, and 1 rRNA that were differentially expressed and confirmed their expression in paired tissues (n = 20) from both phases of the menstrual cycle with the use of qRT-PCR. The results indicated up-regulation of the snoRNAs (SNORD30, SNORD27, SNORA16A, SNORD46, and SNORD56) and down-regulation of the piRNAs (piR-1311, piR-16677, piR-20365, piR-4153), tRNA (TRG-GCC5-1), and rRNA (RNA5SP202) expression in leiomyoma compared with myometrium (P<.05). The pattern of expression of these sncRNAs was similar to RNA sequencing analysis, with no menstrual cycle-dependent differences detected except for SNORD30. Because Argonaute 2 (AGO2) is required for sncRNA-mediated gene silencing, we determined its expression and found greater abundance in leiomyoma.
Conclusion(S):
Our results provide the first evidence for the differential expression of additional classes of sncRNAs and AGO2 in leiomyoma, implicating their roles as a gene regulatory mechanism.
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