Oncotarget

Research Papers:

Fixed differences in the 3'UTR of buffalo PRNP gene provide binding sites for miRNAs post-transcriptional regulation

Hui Zhao, Siqi Wang, Lixia Guo, Yanli Du, Linlin Liu, Tengfei Ma, Newton O. Otecko, Canpeng Li _ and Yaping Zhang

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Oncotarget. 2017; 8:46006-46019. https://doi.org/10.18632/oncotarget.17545

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Abstract

Hui Zhao1,2,*, Siqi Wang1,3,*, Lixia Guo1,3,*, Yanli Du1,3, Linlin Liu1,3, Tengfei Ma1,3, Newton O. Otecko4,5, Canpeng Li6 and Yaping Zhang1,4

1State Key Laboratory for Conservation and Utilization of Bio-resource, Yunnan University, Kunming 650091, P.R. China

2Key Laboratory for Animal Genetic Diversity and Evolution of High Education in Yunnan Province, Yunnan University, Kunming 650091, P.R. China

3School of Life Science, Yunnan University, Kunming 650091, P.R. China

4State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650223, P.R. China

5Kunming College of Life Science, University of Chinese Academy of Sciences, Kunming 650204, P.R. China

6School of Chemical Science and Technology, Yunnan University, Kunming 650091, P.R. China

*These authors contributed equally to this work

Correspondence to:

Canpeng Li, email: [email protected]

Yaping Zhang, email: [email protected]

Keywords: microRNA, difference, BSE, PRNP, 3’UTR

Received: December 02, 2016    Accepted: March 30, 2017    Published: May 02, 2017

ABSTRACT

Bovine spongiform encephalopathy, a member of transmissible spongiform encephalopathies, has not been reported in buffaloes, Bubalus bubalis. Prion protein (PrP), encoded by the prion protein gene (PRNP), is fundamental in the pathogenesis of transmissible spongiform encephalopathies. We previously showed that cattle express more PrP proteins but lower PRNP mRNA than buffaloes in several pivotal tissues like the obex. Therefore, we sought to establish whether genetic variability in PRNP 3’UTR, mediated by miRNA down-regulation, causes PrP expression differences between cattle and buffaloes. We annotated the 3’UTR of buffalo PRNP gene by 3’RACE experiment. A total of 92 fixed differences in the complete 3’UTR (~ 3 kb) were identified between 13 cattle and 13 buffaloes. Resequencing of UTR-C (g.786-1436) and UTR-B (g.778-1456) fragments confirmed that all mutations except g.1022T in cattle are fixed differences between 147 cattle and 146 buffaloes. In addition, analysis of the variation of ΔG between cattle and buffalo sequences reveals four remarkable differences. Two buffalo-specific insertion sites (a 28-bp insertion and an AG insertion in buffalo 3’UTR of PRNP g.970-997 and g. 1088-1089, respectively) and two mutants (g. 1007-1008 TG→CC) create compatible binding sites for miRNAs in buffalo 3’UTR. This was validated through luciferase reporter assays which demonstrated that miR-125b-5p, miR-132-3p, miR-145-5p, miR-331-3p, and miR-338-3p directly act on the fixed difference sites in buffalo 3’UTR. Additional expressional analyses show that these five miRNAs are coexpressed with PRNP in bovine obex tissues. Our study reveals a miRNAs regulated mechanism explaining the differences in prion expression between cattle and buffalo.


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