dictyNews Volume 28 Number 16

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dictyNews 
Electronic Edition 
Volume 28, number 16 
June 8, 2007 

Please submit abstracts of your papers as soon as they have been 
accepted for publication by sending them to dicty@northwestern.edu 
or by using the form at 
http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit. 

Back issues of dictyNews, the Dicty Reference database and other 
useful information is available at dictyBase - http://dictybase.org. 

 
========= 
Abstracts 
========= 

Regulation of G-protein-coupled cAMP receptor activation by a   
hydrophobic 
residue in transmembrane helix 3 

 
Minghang Zhang, Mousumi Goswami, Satoshi Sawai, Edward C. Cox, and 
Dale Hereld 

 
Molecular Microbiology, in press 

 
Summary:  cAR1, a G-protein-coupled cAMP receptor, is essential for 
multicellular development of Dictyostelium.  We previously identified a 
cAR1-Ile104 mutant that appeared to be constitutively activated based on 
its constitutive phosphorylation, elevated affinity for cAMP, and 
dominant-negative effects on development as well as specific cAR1   
pathways that are subject to adaptation.  To investigate how Ile104 might   
regulate cAR1 activation, we assessed the consequences of substituting it with   
all other amino acids.  Constitutive phosphorylation of these Ile104 mutants 
varied broadly, suggesting that they are activated to varying extents, 
and was correlated with polarity of the substituting amino acid residue. 
Remarkably, all Ile104 substitutions, except for the most conservative, 
dramatically elevated the receptor's cAMP affinity.  However, only a   
third of the mutants (those with the most polar substitutions) blocked 
development.  These findings are consistent with a model in which polar 
Ile104 substitutions perturb the equilibrium between inactive and active 
cAR1 conformations in favor of the latter.  Based on homology with   
rhodopsin, Ile104 is likely buried within inactive cAR1 and exposed to the   
cytoplasm upon activation.  We propose that the hydrophobic effect normally   
promotes burial of Ile104 and hence cAR1 inactivation, while polar   
substitution of Ile104 mitigates this effect, resulting in activation. 

 
Submitted by: Dale Hereld [hereldd@niaid.nih.gov] 
-------------------------------------------------------------------------------- 

GxcDD, a putative RacGEF, is involved in Dictyostelium development 

 
Subhanjan Mondal, Dhamodharan Neelamegan, Francisco Rivero, 
Angelika. A. Noegel 

 
Institute for Biochemistry I, Medical Faculty, and Center for Molecular 
Medicine, University of Cologne, 50931 Cologne 

 
BMC Cell Biology 

 
Background 
Rho subfamily GTPases are implicated in a large number of actin-related 
processes. They shuttle from an inactive GDP-bound form to an active   
GTP-bound form. This reaction is catalysed by Guanine nucleotide exchange   
factor (GEFs). GTPase activating proteins (GAPs) help the GTPase return to the inactive 
GDP-bound form. The social amoeba Dictyostelium discoideum lacks a   
Rho or Cdc42 ortholog but has several Rac related GTPases. Compared to our   
understanding of the downstream effects of Racs our understanding of upstream   
mechanisms that activate Rac GTPases is relatively poor. 

 
Results 
We report on GxcDD (Guanine exchange factor for Rac GTPases), a   
Dictyostelium RacGEF. GxcDD is a 180-kDa multidomain protein containing a type 3 CH   
domain, two IQ motifs, three PH domains, a RhoGEF domain and an ArfGAP domain. 
Inactivation of the gene results in defective streaming during   
development under different conditions and a delay in developmental timing. The 
characterization of single domains revealed that the CH domain of GxcDD 
functions as a membrane association domain, the RhoGEF domain can   
physically interact with a subset of Rac GTPases, and the ArfGAP-PH tandem   
accumulates in cortical regions of the cell and on phagosomes. Our results also   
suggest that a conformational change may be required for activation of GxcDD,   
which would be important for its downstream signaling. 

 
Conclusions 
The data indicate that GxcDD is involved in proper streaming and 
development. We propose that GxcDD is not only a component of the Rac 
signaling pathway in Dictyostelium, but is also involved in integrating 
different signals. We provide evidence for a Calponin Homology domain 
acting as a membrane association domain. GxcDD can bind to several Rac 
GTPases, but its function as a nucleotide exchange factor needs to be 
studied further. 

 

Submitted by: Angelika. A. Noegel [noegel@uni-koeln.de] 

 

 
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