Thymosin alpha-1 (Tα1) is a molecule retaining pleiotropic effects toward several pathological conditions, especially acting as a modulator of immune response and inflammation. However, molecular mechanisms underlying the peculiar, wide-range activity of Tα1 have not been completely elucidated. To get information on the mechanism of action of Tα1 and to find new potential applications, we performed the analysis of the transcription profile of 8300 genes in human peripheral blood mononuclear cells in vitro treated with Tα1. Together with a variety of genes already known to be modulated by Tα1, our analysis revealed a remarkable number of genes not yet described before to be modulated by this thymic peptide, revealing a complex scenario that underlines the role of this peptide as a regulator of lymphocyte functions. Here, we report the list of the more representative modulated genes and discuss their implications on the modulation of the innate and adaptive responses. Our results open new perspectives on the use of Tα1 for the treatment of diseases associated with immune-disregulation.