GmRFP1 encodes a previously unknown RING-type E3 ubiquitin ligase in Soybean (Glycine max).
RING-finger proteins with E3 ubiquitin ligase activity play important roles in the regulation of plant growth and development. In this study, a cDNA clone encoding a novel RING-finger protein, designated as GmRFP1, was isolated and characterized from soybean. GmRFP1 was an intronless gene encoding a predicted protein product of 392 amino acid residues with a molecular mass of ~43 kDa. The protein contained a RING-H2 motif and an N-terminal transmembrane domain. The transcript was observed in all detected organs and was up-regulated by abscisic acid (ABA) and salt stress, but down-regulated by cold and drought treatments. We further expressed and purified both wild type and mutant version of GmRFP1 in E. coli. In vitro assays showed that the purified GmRFP1 induced the formation of polyubiquitin chains while mutation within the RING-finger region abolished the ubiquitination activity. These findings suggest that GmRFP1 is a previously unknown E3 ubiquitin ligase in soybean and that the RING domain is required for its activity. It may play unappreciated roles in ABA signaling and stress responses via mediating the ubiquitination and degradation of target proteins through the ubiquitin-proteasome pathway.
National Center for Soybean Improvement, National Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, 210095, Nanjing, People's Republic of China.
Qiu-Li
QL
Du
Wen-Zhuo
WZ
Cui
Chun-Hong
CH
Zhang
De-Yue
DY
Yu
D000040:Abscisic Acid / Q000494:pharmacology; D000064:Acclimatization / Q000235:genetics; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003080:Cold Temperature / Q000009:adverse effects; D055864:Droughts; D017343:Genes, Plant / Q000502:physiology; D008969:Molecular Sequence Data; D054829:RING Finger Domains / Q000235:genetics; D011994:Recombinant Proteins / Q000302:isolation & purification; D055049:Salt Tolerance / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013025:Glycine max / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D054875:Ubiquitination / Q000235:genetics
D011994:Recombinant Proteins; D000040:Abscisic Acid; D044767:Ubiquitin-Protein Ligases