| HSP40s are a group of J-domain containing co-chaporone for HSP70,which play important roles in stress responses.We previously reported that over-expression of a soybean Gm HSP40.1 in Arabidposis induced cell death and activated defense response.We subsequently found that over-expressing Gm HSP40.1 could also result in a significantly delayed flowering phenotype in Arabidopsis in a day-length independent manner,indicating a pleiotropic effects of the Gm HSP40.1over-expression.It has been reported that EIP9 participates in flowering regulation through interacting with EMF1(EMBRYONIC FLOWER 1).EMF1 is one the the most important components of the POLYCOMB GROUP COMPLEX 1(PRC1),which plays a critical role in histone methylations.Even though the EIP9-EMF1 interaction has been shown in the previous report,the functional importance of this interaction in regulation of flowering has not been further investigated.EMF1c(EMF1 complex)is a newly identified complex,in which EMF1,LHP1 and JMJ14 are the major components.EMF1 c is responsible for the histone modifications for the gene loci that control flowering time,especially FT.Our Y2 H assays indicated that Gm HSP40.1 interacted with both EMF1 and JMJ14,but not LHP1;The interaction in the nuclei between Gm HSP40.1 and JMJ14 was further confirmed by a Bi FC assay,suggesting that Gm HSP40.1 may exert its function on flowering through interacting with EMF1 and JMJ14,the major components of the EMF1 c.Genetic analysis indicated that EMF1 was epistatic to Gm HSP40.1,and the loss-of-function mutation in JMJ14 could partially rescue the late flowering phenotype of Gm HSP40.1-OE line,further confirming the functional connections among these three proteins.Thus it is reasonable to speculate that Gm HSP40.1 participates in the regulation of flowering time by modulating EMF1 c function through interacting with both EMF1 and JMJ14.The fact that the eip9-2 partially rescued the late flowering phenotype indicated that Gm HSP40.1 and EIP9 are functional equivalent.Ch IP-q PCR results showed that,while the level of H3K27me3 modification was significantly increased in the promoter region of FT gene,the level of H3K4me3 modification was significantly at-1200 bp,-1000 bp promoter regions and 150 bp before second exon regions of FT gene.On the contrary,the H3K4me3 and H3K27me3 modifications were significantly increased and decreased,respectively,at FLC locus.Consistent with the late flowering phenotype and Ch IP-q PCR results,the expression levels of the genes encoding the positive and negative regulators of flowering time were significantly increased and decreased,respectively,in the Gm HSP40.1-OE lines.Furthermore,confocol experiments showed that both Gm HSP40.1,EMF1 and JMJ14 localized in the nucleus.Together,our results indicate that Gm HSP40.1 participates in the regulation of flowering time by modulating EMF1 c function through interacting with both EMF1 and JMJ14. |
