A novel family of cysteine-rich secreted proteins with unique tissue distribution has recently been identified. One of the members, resistin (for "resistance to insulin"), also called FIZZ3, was identified in a screen for molecules that are down-regulated in mature adipocytes upon administration of thiazolidinediones. The prototypical member of this family was originally identified from bronchoalveolar lavage fluid of inflamed lungs and designated FIZZ1 ("found in inflammatory zone"). This molecule was also found to be highly expressed in adipose tissue and was named resistin-like molecule alpha (RELMalpha). Here we demonstrate that RELMalpha inhibits the differentiation of 3T3-L1 preadipocytes into adipocytes. RELMalpha has no effect on proliferation of 3T3-L1 preadipocytes. Pretreatment of 3T3-L1 preadipocytes with RELMalpha does not affect insulin- or platelet-derived growth factor-induced mitogenesis. IRS-1 phosphorylation and glucose transport stimulated by insulin in mature adipocytes were also unaffected by RELMalpha. We show that RELMalpha forms disulfide-linked homooligomers based on results from electrophoresis under reducing and nonreducing conditions, coimmunoprecipitation experiments as well as by mass spectrometry. In addition, RELMalpha is able to form heterooligomers with resistin but not RELMbeta. Since RELMalpha is expressed by adipose tissue and it is a secreted factor, our findings suggest that RELMalpha may be involved in the control of the adipogenesis as well as in the process of muscle differentiation.