In contrast to the molecular etiology of autonomously functioning thyroid nodules, the molecular cause of cold thyroid nodules (CTNs), their benign, functional inactive counterparts, are so far largely unknown. Because of the partially dedifferentiated phenotype of CTNs, alterations in signaling cascades that favor proliferation, but not differentiation, are likely candidates for tumor induction and progression. The importance of RAS mutations for the development of benign nodules with follicular histology is still in question. However, differentially expressed genes in the context of their signaling cascades could define aberrant signaling in CTNs. Therefore, we investigated gene expression in 22 CTNs and their normal surrounding tissue using Affymetrix GeneChips. Most prominently, data analysis revealed an increased expression of cell cycle-associated genes and a special relevance of protein kinase C signaling, whereas no evidence of RAS-MAPK signaling in CTNs was found. Moreover, we determined 31 differentially regulated genes in CTNs, including several histone mRNAs. Taken together, these results explain recent findings showing an increased proliferation in CTNs and draw attention to protein kinase C signaling, but away from RAS-MAPK signaling, as being involved in the etiology of CTNs.