Transcriptional Regulation Mechanism Of RhERF001 Involved In Petal Abscission In Rose

Rose is one of the most important cut flowers in world.The sale share of rose cut flower accounts for about 33.3%of annual cut flower trade in the world.Premature rose petal shedding results in significant shortening of the ornamental period and ornamental value.Therefore,developing freshness agent that delays or prevents abscission,and making cultivars that are not prone to abscission through molecular breeding are the long-term objectives for the researcher.It is well-known that the abscission of petal was induced by ethylene,but is unclear that the physiological mechanism of hormones regulate petal abscission.In this study,we employed the abscission-prone(AP)rose cultivar 'Gloden shower',abscission-resistant(AR)rose cultivars 'Gold medal' and 'Samantha' as the research materials,explored biological function and regulatory mechanism of an ethylene signaling component-RhERF001 in rose petal abscission.Main results are as follows:(1)We found that the cellular morphology was changing during abscission process in rose abscission zone(AZ)by anatomical analysis of AP rose 'Golden shower',AR rose 'Gold medal'and 'Samantha'.Generally,the cells were roundness in stage 0,rectangle in stage 2,and then cells became to roundness because of the degradation of cell walls and middle layers after shedding occurred in three rose cultivars.In addition,we found the timing of AZ development was different among three rose cultivars.The AZ of AP rose cultivar 'Golden shower' shed in stage 3,although the AZ of AR rose cultivars 'Gold medal'abscised in stage 5,and the AR rose cultivars 'Samantha' not shed in stge 6.Therefore,we deduced that AZ may activate in stage 4,and the AZ may separate after stage 6 in the AR rose cultivars 'Samantha'.(2)We obtained many abscission-related genes by RNA-seq using 'Golden shower' and 'Gold Medal',including RSA41424,which had high expression level in stage 1 to stage 3,and quickly reduced in stage 4,then almost disappeared in the following stages in rose petal AZ tissue.RSA41424 was named as RhERF001 though blast analysis.(3)We investigated the biological function of RhERF001 in rose petal abscission using virus-induced gene silencing(VIGS)in 'Gold Medal' flowers and 'Samantha' seeding.In the RhERF001-silencing'Gold Medal' flowers,no difference was observed on flower opening compared to TRV2 control flowers,but the petal abscission of RhERF001-silencing flowers were accelerated 2 days compared to petal abscission of control,the average abscission time is 9 days after infected in TRV2 control flowers,but 7 days in RhERF001-silencing flowers,shortened 2 days.In the RhERF001-silencing 'Samantha' plantlets,the average abscission time was 10 days after flower fully opening in TRV2 control flowers,but 6 days in RhERF001-silencing flowers,shortened 4 days.It suggested that the abscission of rose petal was suppressed by RhERF001.(4)We obtained 1400 differential expression genes by RNA-Seq using RhERF001-silencing'Samantha'plantlets.The differential expression genes included 55 transcription factors,64 protein kinases,and 982 functional proteins.We selected the transcription factor RhERF4 and the functional protein RhBGLA1,and verified of the combination of upstream and downstream by promoter analysis including yeast one-hybrid and dual-luciferase(?)reporter assay systems.The results showed that RhERF4 was a downstream gene of the RhERF001,and the directly upstream gene of RhBGLA1.Furthermore,we observed the content of pectin and the permeability of vascular bundle in petal AZ of RhERF001-silencing 'Samantha'plantlets.We found that the content of demethyl-pectin was less and the vascular bundle was more blocked in AZ of RhERF001-silencing flowers.The results suggested that RhERF001 regulates the rose petal abscission by altering the content of demethyl-pectin and the permeability of vascular bundle.In conclusion,RhERF001 is a repressor involved in rose petal abscission.The expression of RhERF001 was significantly decresed after abscission initiation,and then RhERF001 enhanced the expression of RhBGLA1 by a transcription repressor RhERF4,eventually accelerated the petal abscission by promoted the degradation of pectin.