Among the members of the 5'-nucleotidase family, there is only one membrane-bound ectosolic isoenzyme. This esterase prefers AMP as substrate but can hydrolyze a number of purine and pyrimidine phosphorylated compounds, indicating that no evolutive pressure to develop a more restricted specificity was exerted on this enzyme. On the contrary, five cytosolic isoforms have been evolved, probably by convergent evolution, showing different and restricted substrate specificity. The different isoforms have different level of expression and distribution in organs of vertebrates. The cytosolic nucleotidase specific for IMP and GMP (cN-II), is an enzyme allosterically regulated, structurally strongly conserved and expressed at a low but constant level in all organs and tissues in vertebrates. As far as we know, alteration of cN-II expression is limited to pathological conditions. In this review, we report the results of the modulation of cN-II specific activity exerted by silencing or hyperexpression in different cell types, in the attempt to better understand its role and implications in pathology and therapy.