75 Integrating Membrane Transport with Male Gametophyte ... - TAIR

385 Identification and Characterization of a Novel, Recessive Allele of CUL1 that Disrupts SCF
regulation at the C-terminus of CUL1
Jonathan Gilkerson 1, 2 , Jessica Brown 2 , Judy Callis 2, 1
1
UC Davis, Plant Biology Graduate Group, 2 UC Davis, Section of Molecular and Cellular Biology
Auxin regulates many aspects of plant growth and development and has been shown to signal via selected proteolysis
of AUX/IAA proteins. Auxin facilitates the interaction between the AUX/IAAs and an SCF E3 ubiquitin ligase. SCFtype
E3s are the most abundant type of ubiquitin ligase in Arabidopsis and several other signaling pathways including
those for jasmonic acid, gibberellic acid, and the circadian clock are mediated by SCF-mediated protein degradation.
The SCF ubiquitin ligase is a multi-subunit protein complex consisting of four subunits: SKP1 (ASK1 in Arabidopsis),
a Cullin family member such as CUL1, a RING protein RBX1, and a target recognizing F-box protein. Here, we report
the characterization of a novel, recessive allele of CUL1, cul1-6, isolated from a genetic screen in Arabidopsis thaliana
(Col) designed to identify plants defective in degradation of a fusion protein with an Aux/IAA degron, IAA1:LUC. One
mutation was identified that exhibited very slow IAA1:LUC degradation compared to the 10-12 min half-life of IAA1:
LUC in the progenitor line as measured in cycloheximide chases. Genetic mapping placed the mutation within a 0.8 cM
genetic interval that includes the CULLIN1 gene. We sequenced the coding region, 5’ UTR, and 3’UTR of CUL1 and
identified a mutation. Aerial portions of cul1-6 exhibit reduced apical dominance, delayed senescence, reduced fertility,
and small, wrinkled rosette leaves. In root growth assays, the mutant roots were shorter, had fewer lateral roots than
wild type, and were less responsive to 2,4-D. Genetic complementation by expressing FLAG-tagged wild type CUL1
under control of its endogenous promoter confirmed that this locus was responsible for the mutant phenotype. Mutant
alleles of CUL1 have been isolated previously (Development 127:23-32; Plant J 43:371-83), but the mutations are all
at the N-terminus and affect binding to the SCF subunit SKP1. No alleles that affect subunit interactions at the CUL1
C-terminus have been described to date. We believe the location of the mutation within CUL1 and its recessive nature
will allow this allele to be used as tool to dissect regulation of SCF activity at the C-terminus of CUL1. This work was
supported in part by National Science Foundation IBN 0212659 to JC.
386 SPINDLY-Dependant, DELLA-Independent Gibberellin Signaling Pathway to Suppress
Cytokinin Responses
Yaarit Greenboim-Wainberg, David Weiss
The Robert H. Smith Institute of Plant Sciences and Genetics in Agriculture, Faculty of Agricultural, Food and
Environmental Quality Sciences, The Hebrew University of Jerusalem, Rehovot, Israel
We have shown previously that SPINDLY (SPY), a negative regulator of GA (gibberellin) signaling, promotes
cytokinin responses in Arabidopsis. In addition, GA also represses the effects of cytokinin, suggesting that there is cross
talk between the two hormone-response pathways and that this crosstalk requires SPY function. We demonstrated that
SPY acts as both a repressor of GA responses and a positive regulator of cytokinin signaling and suggested that GA
suppresses cytokinin responses through the inhibition of SPY. The DELLA proteins play a key role in the regulation of
GA-signal transduction and in the interactions between GA and auxin, ethylene or ABA response pathways. Here, we
determine whether the DELLA proteins also regulate the interaction between the gibberellin and cytokinin signaling
pathways. GA-regulation of inflorescence-stem elongation depends on DELLA (GAI and RGA) protein activities and
GA inhibits the suppressed inflorescence stem elongation after application of cytokinin. Cytokinin may arrest elongation
by increasing the stability of the DELLA proteins, and GA may inhibit this effect by promoting the latter's degradation.
Our results show that cytokinin has no effect on RGA stability or on GA-induced RGA degradation. Furthermore, the
inhibition of stem elongation by cytokinin was suppressed in spy plants but not by the loss of GAI and RGA activities,
suggesting that the GA signal generated by the loss of these DELLA proteins does not suppress cytokinin responses.
Cytokinin also inhibited inflorescence elongation in the dwarf, gain-of-function gai. While GA's effect on stem elongation
was completely blocked in gai plants, GA could still act in the mutant's stem to inhibit cytokinin-suppressed elongation.
We suggest that the GA signal to suppress cytokinin responses is mediated by SPY but not by the DELLA proteins, and
therefore it is possible that SPY acts in a DELLA-independent GA signaling pathway.