Molecular Coevolution of Nuclear and Nucleolar Localization Signals inside the Basic Domain of HIV-1 Tat.

During evolution, viruses had to adapt to an increasingly complex environment of eukaryotic cells. Viral proteins that need to enter the cell nucleus or associate with nucleoli possess nuclear localization signals (NLSs) and nucleolar localization signals (NoLSs) for nuclear and nucleolar accumulation, respectively. As viral proteins are relatively small, acquisition of novel sequences seems to be a more complicated task for viruses than for eukaryotes. Here, we carried out a comprehensive analysis of the basic domain (BD) of HIV-1 Tat to show how viral proteins might evolve with NLSs and NoLSs without an increase in protein size. The HIV-1 Tat BD is involved in several functions, the most important being the transactivation of viral transcription. The BD also functions as an NLS, although it is substantially longer than a typical NLS. It seems that different regions in the BD could function as NLSs due to its enrichment with positively charged amino acids. Additionally, the high positive net charge inevitably causes the BD to function as an NoLS through a charge-specific mechanism. The integration of NLSs and NoLSs into functional domains enriched with positively charged amino acids might be a mechanism that allows the condensation of different functional sequences in small protein regions and, as a result, reduces protein size, influencing the origin and evolution of NLSs and NoLSs in viruses. IMPORTANCE Here, we investigated the molecular mechanism of nuclear localization signal (NLS) and nucleolar localization signal (NoLS) integration into the basic domain of HIV-1 Tat (49RKKRRQRRR57) and found that these two supplementary functions (i.e., function of NLS and function of NoLS) are embedded in the basic domain amino acid sequence. The integration of NLSs and NoLSs into functional domains of viral proteins enriched with positively charged amino acids is a mechanism that allows the concentration of different functions within small protein regions. Integration of NLS and NoLS into functional protein domains might have influenced the viral evolution, as this could prevent an increase in the protein size.

The GAR domain integrates functions that are necessary for the proper localization of fibrillarin (FBL) inside eukaryotic cells.

Fibrillarin (FBL) is an essential nucleolar protein that participates in pre-rRNA methylation and processing. The methyltransferase domain of FBL is an example of an extremely well-conserved protein domain in which the amino acid sequence was not substantially modified during the evolution from Archaea to Eukaryota. An additional N-terminal glycine-arginine-rich (GAR) domain is present in the FBL of eukaryotes. Here, we demonstrate that the GAR domain is involved in FBL functioning and integrates the functions of the nuclear localization signal and the nucleolar localization signal (NoLS). The methylation of the arginine residues in the GAR domain is necessary for nuclear import but decreases the efficiency of nucleolar retention via the NoLS. The presented data indicate that the GAR domain can be considered an evolutionary innovation that integrates several functional activities and thereby adapts FBL to the highly compartmentalized content of the eukaryotic cell.

Origin of the nuclear proteome on the basis of pre-existing nuclear localization signals in prokaryotic proteins.

BACKGROUND: The origin of the selective nuclear protein import machinery, which consists of nuclear pore complexes and adaptor molecules interacting with the nuclear localization signals (NLSs) of cargo molecules, is one of the most important events in the evolution of eukaryotic cells. How proteins were selected for import into the forming nucleus remains an open question. RESULTS: Here, we demonstrate that functional NLSs may be integrated in the nucleotide-binding domains of both eukaryotic and prokaryotic proteins and may coevolve with these domains. CONCLUSION: The presence of sequences similar to NLSs in the DNA-binding domains of prokaryotic proteins might have created an advantage for nuclear accumulation of these proteins during evolution of the nuclear-cytoplasmic barrier, influencing which proteins accumulated and became compartmentalized inside the forming nucleus (i.e., the content of the nuclear proteome). REVIEWERS: This article was reviewed by Sergey Melnikov and Igor Rogozin. OPEN PEER REVIEW: Reviewed by Sergey Melnikov and Igor Rogozin. For the full reviews, please go to the Reviewers' comments section.

Switching of cardiac troponin I between nuclear and cytoplasmic localization during muscle differentiation.

The nuclear accumulation of proteins may depend on the presence of short targeting sequences, which are known as nuclear localization signals (NLSs). Here, we found that NLSs are predicted in some cytosolic proteins and examined the hypothesis that these NLSs may be functional under certain conditions. As a model, human cardiac troponin I (hcTnI) was used. After expression in cultured non-muscle or undifferentiated muscle cells, hcTnI accumulated inside nuclei. Several NLSs were predicted and confirmed by site-directed mutagenesis in hcTnI. Nuclear import occurred via the classical karyopherin-α/ß nuclear import pathway. However, hcTnI expressed in cultured myoblasts redistributed from the nucleus to the cytoplasm, where it was integrated into forming myofibrils after the induction of muscle differentiation. It appears that the dynamic retention of proteins inside cytoplasmic structures can lead to switching between nuclear and cytoplasmic localization.

Proliferation, cancer, and aging-novel functions of the nucleolar methyltransferase fibrillarin?

Fibrillarin is an essential nucleolar protein that catalyzes the 2'-O-methylation of ribosomal RNAs. Recently, experimental data have begun to accumulate that suggest that fibrillarin can influence various cellular processes, development of pathological processes, and even aging. The exact mechanism by which fibrillarin can influence these processes has not been found, but some experimental data indicate that up- or downregulation of fibrillarin can modify the ribosome structure and, thus, causе an alteration in relative efficiency with which various mRNAs are translated. Here, we discuss recent studies on the potential roles of fibrillarin in the regulation of cell proliferation, cancer progression, and aging.