Rat Nopp140, a nonribosomal protein of the nucleolus and coiled bodies, was characterized as one of the most highly phosphorylated proteins in the cell. Based on its subcellular location, its nuclear localization signal binding capacity, and its shuttling between the nucleolus and the cytoplasm, Nopp140 was proposed to function as a chaperone in ribosome biogenesis. This study shows that casein kinase II phosphorylates Nopp140 to its unusual high degree and identifies the yeast SRP40 gene product as immunologically and structurally related to rat Nopp140. SRP40 encodes an acidic (pI = 3. 9), serine-rich (49%) protein of 41 kDa whose carboxyl terminus exhibits 59% sequence identity to that of Nopp140. SRP40 localizes to the yeast nucleolus and is required at a specific cellular concentration for optimal growth as indicated by the negative effect on cell growth of both overexpression and deletion of its gene. Like Nopp140, SRP40 is phosphorylated by casein kinase II, but to a much lesser extent. While the parallels between these two proteins suggest that SRP40 is the bona fide yeast Nopp140 homolog, their disparities reflect the differences in nucleolar dynamics and regulation of ribosome biogenesis between yeast and vertebrates.