Abstract
miR-29a is a conserved miRNA that participates in bone formation and immune response in vertebrates. miR-29a of Pinctada martensii (Pm-miR-29a) was identified in the previous research though deep sequencing. In this report, the precise sequence of mature Pm-miR-29a was validated using miRNA rapid amplification of cDNA ends (miR-RACE) technology. The precursor sequence of Pm-miR-29a was predicted to have 87 bp. Stem loop qRT-PCR analysis showed that Pm-miR-29a was easily detected in all the tissues, although expressions in the mantle and gill were low. The microstructure showed the disrupted growth of the nacre after Pm-miR-29a over-expression, which was induced by mimic injection into P. martensii. Results of the target analysis indicated that neuropeptide Y receptor type 2 (Y2R) was the potential target of Pm-miR-29a. Meanwhile, Pm-miR-29a mimics could obviously inhibit the relative luciferase activity of the reporter containing 3′ UTR (Untranslated Regions) of the Y2R gene. Furthermore, the expression of Y2R was downregulated whereas expressions of interleukin 17 (IL-17) and nuclear factor κB (NF-κB) were upregulated after Pm-miR-29a over-expression in the mantle and gill, thereby suggesting that Pm-miR-29a could activate the immune response of the pearl oyster. Results showed that Pm-miR-29a was involved in nacre formation and immune response by regulating Y2R in pearl oyster P. martensii.
Highlights
MicroRNAs are a kind of endogenous non-coding small single-stranded RNA (18–25 nucleotides)
NNoottaabbllyy,aafftteerrPPmm--mmiiRR--2299aaoovveerr--eexxpprreessssiioonniinnPP..mmaarrtteennssii,i, ggrroowwtthhoofftthheennaaccrreeininththeesshheelllwwaassddisisrruuppteteddaannddaahhoonneeyyccoommbbggrroowwththppaatteterrnnwwaassoobbsseerrvveedd. .TThhiiss pphheennoommeennoonnininddicicaateteddththaattPPmm-m-miRiR-2-299aaccoouuldldreregguulalatetennaaccrerefoformrmaatitoionn. .BBaasseeddoonntthheeffuunnccttiioonnooff mmiRiR--2299aaiinnmmaammmmaalsls,wweepprrooppoosseeddtthhaatttthheeddiissrruupptteeddggrroowwtthhaafftteerrPPmm--mmiiRR--2299aaoovveerr--eexxpprreessssioionnisis possibly caused by the inhibited differentiation of the calcified cells or by the reduced expression of some genes related to biomineralization
Our analysis showed that Pm-miR-29a over-expression could increase the expression of PfIL-17 and nuclear factor κB (NF-κB), indicating that Pm-miR-29a could activate the P. martensii immune response against intracellular pathogens
Summary
MicroRNAs (miRNAs) are a kind of endogenous non-coding small single-stranded RNA (18–25 nucleotides). Increasing evidence has shown that miRNA participates in most cellular processes, such as cell proliferation, differentiation, apoptosis, and immunity, by targeting various genes [2,3,4]. .BBaasseeddoonntthheeffuunnccttiioonnooff mmiRiR--2299aaiinnmmaammmmaalsls,,wweepprrooppoosseeddtthhaatttthheeddiissrruupptteeddggrroowwtthhaafftteerrPPmm--mmiiRR--2299aaoovveerr--eexxpprreessssioionnisis possibly caused by the inhibited differentiation of the calcified cells or by the reduced expression of some genes related to biomineralization. Over-expression of miR-29a suppressed the immune responses induced by intracellular pathogens by targeting IFN-γ [20] and decreasing IL-6 [21] and TNF-α [21] secretions. The different functions of miR-29a in the immune response of mammals and P. martensii may be induced by the different target genes, thereby suggesting that the function of miRNA may change during evolution. Our results suggested that Pm-miR-29a participates in nacre formation and the immune response of P. martensii by targeting Y2R
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