Functional Analysis Of GhNAC12, A Leaf Senescence-related Gene In Cotton (Gossypium Hirsutum L.)

As a kind of programed cell death (PCD), senescence is the last stage of plant development, which is anorderly self-destruction process under the control of nucleus and regulated by diverse environmentalsignals. The previous research in Arabidopsis showed the importance of transcription factors in theregulation of leaf senescence. The NAC is one of the biggest transcription factor families in plant andthere are many leaf senescence-related NAC genes cloned from Arabidopsis and rice, which implies thatthe function analysis of NAC genes is essential to explore the molecular mechanism of leaf senescence.The NAC was named because of the NAC domain consisting of～150highly conserved amino acids inthe N-terminal. NAC transcription factors are widely involved in plant growth, development andresponse to stresses. In this work, GhNAC12was selected, a NAC gene cloned by our lab, to performthe function analysis through the in vitro transcription activation assay, qRT-PCR and transgenicverification, which provides foundation for the research on the molecular mechanism of early-aging incotton. Our results are mainly as follows:1. We cloned a1305-bp genomic CDS sequence of GhNAC12from CCRI10. Sequence alignmentindicated that it had three exons and two introns and both the introns were characterized bycommon features of introns in plants.2. A1592-bp upstream regulatory region was cloned from CCRI10. The cis-acting regulatory elementanalysis showed that there were many elements responding to phytohormone and stress, whichindicated that GhNAC12is involved in plant growth, development and response to stresses.3. The domain analysis suggested that GhNAC12belonged to the NAM subfamily.4. The in vitro transcription activation assay implied that GhNAC12had the transcription activationactivity and could activate the expression of the downstream gene.5. The qRT-PCR data of different varieties showed that the expression level was significant higher inthe CCRI10than that in the Liao4086, which implied that GhNAC12was a candidate gene leadingearly-mature variety to early-aging in upland cotton.6. Overexpression of GhNAC12in Arabidopsis leaded to early senescence.7. qRT-PCR data of roots infected by VD080showed that GhNAC12exhibited a periodic expressionpattern in the root, which suggested that GhNAC12may be involved in the intrusion prevention ofVD080into root in cotton.