title: A charge-dependent mechanism is responsible for the dynamic accumulation of proteins inside nucleoli.
abstract: The majority of known nucleolar proteins are freely exchanged between the nucleolus and the surrounding nucleoplasm. One way proteins are retained in the nucleoli is by the presence of specific amino acid sequences, namely nucleolar localization signals (NoLSs). The mechanism by which NoLSs retain proteins inside the nucleoli is still unclear. Here, we present data showing that the charge-dependent (electrostatic) interactions of NoLSs with nucleolar components lead to nucleolar accumulation as follows: (i) known NoLSs are enriched in positively charged amino acids, but the NoLS structure is highly heterogeneous, and it is not possible to identify a consensus sequence for this type of signal; (ii) in two analyzed proteins (NF-κB-inducing kinase and HIV-1 Tat), the NoLS corresponds to a region that is enriched for positively charged amino acid residues; substituting charged amino acids with non-charged ones reduced the nucleolar accumulation in proportion to the charge reduction, and nucleolar accumulation efficiency was strongly correlated with the predicted charge of the tested sequences; and (iii) sequences containing only lysine or arginine residues (which were referred to as imitative NoLSs, or iNoLSs) are accumulated in the nucleoli in a charge-dependent manner. The results of experiments with iNoLSs suggested that charge-dependent accumulation inside the nucleoli was dependent on interactions with nucleolar RNAs. The results of this work are consistent with the hypothesis that nucleolar protein accumulation by NoLSs can be determined by the electrostatic interaction of positively charged regions with nucleolar RNAs rather than by any sequence-specific mechanism.
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119991, Russia.
authors.forename: Yana R
authors.initials: YR
authors.lastname: Musinova
authors.affiliation: Department of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow 119991, Russia.
authors.forename: Eugenia Y
authors.initials: EY
authors.lastname: Kananykhina
authors.affiliation: Department of Cell Biology and Histology, Lomonosov Moscow State University, Moscow 119991, Russia.
authors.forename: Daria M
authors.initials: DM
authors.lastname: Potashnikova
authors.affiliation: Department of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow 119991, Russia.
authors.forename: Olga M
authors.initials: OM
authors.lastname: Lisitsyna
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119991, Russia. Electronic address: sheval_e@belozersky.msu.ru.
authors.forename: Eugene V
authors.initials: EV
authors.lastname: Sheval
mesh_terms: D000595:Amino Acid Sequence; D002466:Cell Nucleolus / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism*
chemical_list: D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins
keywords: B23; Nucleolar localization signal; Nucleolus; RNA