A minor population of dendritic epidermal T cells (DETC) express the Vγ1. 1Cγ4Vδ6 T cell receptor and T cell clones and hybridomas derived from this subset constitutively secrete cytokines in culture secondary to recognition of an autoantigen. Activation of these autoreactive cells requires the use of the vitronection receptor (VNR) as an accessory molecule which interacts with the Arg‐Gly‐Asp‐Ser (RGDS) sequence of extracellular matrix (ECM) proteins. We have compared the functional properties of Cγ4+ hybridomas derived from newborn thymocytes and from adult spleen with the DETC hybridomas/lines in terms of their ability to secrete cytokines spontaneously and for the use of the VNR as an accessory molecule. Almost all of the Cγ4+ thymocyte hybridomas secreted cytokines spontaneously and in the majority of lines the most prominent cytokine secreted was granulocyte‐monocyte colony‐stimulating factor. In contrast, none of the four splenic Cγ4+ hybridomas secreted cytokines spontaneously although all were capable of cytokine production following activation via the T cell receptor. Although the thymocyte hybridomas did not grow as adherent cell lines in culture, constitutive cytokine production required engagement of the VNR by its ligand in ECM proteins. In all cases, cytokine production could be inhibited by an anti‐VNR monoclonal antibody as well as by soluble RGDS. The strong correlation of functional and molecular properties (see accompanying report) between thymocyte Cγ4+ hybridomas and Cγ4+ DETC suggests that the Cγ4+ DETC may be of thymic origin and that cells with potential for autoreactivity residing in the thymus at birth may populate other peripheral locations in the mouse. The data also support the concept that the VNR, and possibly other integrins, play a role as accessory elements for autoreactive cells and may be essential for the regulation of such activity.