Inhibition of Rho at different stages of thymocyte development gives different perspectives on Rho function.
Development of thymocytes can be staged according to the levels of expression of the cell-surface markers CD4, CD8, CD44, CD25 and CD2. Thymocyte development is regulated by a complex signalling network [1], one component of which is the GTPase Rho. The bacterial enzyme C3 transferase from Clostridium botulinum selectively ADP-ribosylates Rho in its effector-binding domain and thereby abolishes its biological function [2,3]. To explore the function of Rho in thymocyte development, we previously used the proximal promoter of the gene encoding the Src-family kinase p56lck to make transgenic mice that selectively express C3 transferase in the thymus [4,6]. In these mice, which lack Rho function from the earliest thymocyte stages, thymocyte numbers are reduced by approximately 50- to 100-fold. Here, we describe transgenic mice that express C3 transferase under the control of the locus control region (LCR) of the CD2 gene; this regulatory element drives expression at a later stage of thymocyte development than the lck proximal promoter [7]. In these mice, thymocyte numbers were also reduced by 50- to 100-fold, but unlike the lck-C3 mice, in which the reduction predominantly results from defects in cell survival of CD25(+) thymocyte progenitors, the CD2-C3 transgenic mice had a pre-T-cell differentiation block at the CD25(+) stage after rearrangement of the T-cell receptor (TCR) beta chains. Analysis of CD2-C3 mice demonstrated that Rho acts as an intracellular switch for TCR beta selection, the critical thymic-differentiation checkpoint. These results show that Rho-mediated survival signals for CD25(+) pre-T cells are generated by the extracellular signals that act on earlier thymocyte precursors and also that temporal cell-type-specific elimination of Rho can reveal different functions of this GTPase in vivo.
Cleverley
S
S
Lymphocyte Activation Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London, WC2A 3PX, UK. s.cleverley@icrf.icnet.uk
Henning
S
S
Cantrell
D
D
D036002:ADP Ribose Transferases / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001905:Botulinum Toxins; D018801:CD2 Antigens / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D003014:Clostridium botulinum / Q000201:enzymology / Q000235:genetics; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D005798:Genes, Bacterial; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics; D013601:T-Lymphocytes / Q000166:cytology* / Q000276:immunology / Q000378:metabolism*
D018801:CD2 Antigens; D020690:GTPase-Activating Proteins; D016693:Receptors, Antigen, T-Cell, alpha-beta; C085379:rho GTPase-activating protein; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D001905:Botulinum Toxins; D019204:GTP-Binding Proteins