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      Functional equivalence of germ plasm organizers

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          Abstract

          The proteins Oskar (Osk) in Drosophila and Bucky ball (Buc) in zebrafish act as germ plasm organizers. Both proteins recapitulate germ plasm activities but seem to be unique to their animal groups. Here, we discover that Osk and Buc show similar activities during germ cell specification. Drosophila Osk induces additional PGCs in zebrafish. Surprisingly, Osk and Buc do not show homologous protein motifs that would explain their related function. Nonetheless, we detect that both proteins contain stretches of intrinsically disordered regions (IDRs), which seem to be involved in protein aggregation. IDRs are known to rapidly change their sequence during evolution, which might obscure biochemical interaction motifs. Indeed, we show that Buc binds to the known Oskar interactors Vasa protein and nanos mRNA indicating conserved biochemical activities. These data provide a molecular framework for two proteins with unrelated sequence but with equivalent function to assemble a conserved core-complex nucleating germ plasm.

          Author summary

          Multicellular organisms use gametes for their propagation. Gametes are formed from germ cells, which are specified during embryogenesis in some animals by the inheritance of RNP granules known as germ plasm. Transplantation of germ plasm induces extra germ cells, whereas germ plasm ablation leads to the loss of gametes and sterility. Therefore, germ plasm is key for germ cell formation and reproduction. However, the molecular mechanisms of germ cell specification by germ plasm in the vertebrate embryo remain an unsolved question. Proteins, which assemble the germ plasm, are known as germ plasm organizers. Here, we show that the two germ plasm organizers Oskar from the fly and Bucky ball from the fish show similar functions by using a cross species approach. Both are intrinsically disordered proteins, which rapidly changed their sequence during evolution. Moreover, both proteins still interact with conserved components of the germ cell specification pathway. These data might provide a first example of two proteins with the same biological role, but distinct sequence.

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          Visualization of interactions among bZIP and Rel family proteins in living cells using bimolecular fluorescence complementation.

          Networks of protein interactions coordinate cellular functions. We describe a bimolecular fluorescence complementation (BiFC) assay for determination of the locations of protein interactions in living cells. This approach is based on complementation between two nonfluorescent fragments of the yellow fluorescent protein (YFP) when they are brought together by interactions between proteins fused to each fragment. BiFC analysis was used to investigate interactions among bZIP and Rel family transcription factors. Regions outside the bZIP domains determined the locations of bZIP protein interactions. The subcellular sites of protein interactions were regulated by signaling. Cross-family interactions between bZIP and Rel proteins affected their subcellular localization and modulated transcription activation. These results attest to the general applicability of the BiFC assay for studies of protein interactions.
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            A role for Piwi and piRNAs in germ cell maintenance and transposon silencing in Zebrafish.

            Piwi proteins specify an animal-specific subclass of the Argonaute family that, in vertebrates, is specifically expressed in germ cells. We demonstrate that zebrafish Piwi (Ziwi) is expressed in both the male and the female gonad and is a component of a germline-specifying structure called nuage. Loss of Ziwi function results in a progressive loss of germ cells due to apoptosis during larval development. In animals that have reduced Ziwi function, germ cells are maintained but display abnormal levels of apoptosis in adults. In mammals, Piwi proteins associate with approximately 29-nucleotide-long, testis-specific RNA molecules called piRNAs. Here we show that zebrafish piRNAs are present in both ovary and testis. Many of these are derived from transposons, implicating a role for piRNAs in the silencing of repetitive elements in vertebrates. Furthermore, we show that piRNAs are Dicer independent and that their 3' end likely carries a 2'O-Methyl modification.
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              Length-dependent prediction of protein intrinsic disorder

              Background Due to the functional importance of intrinsically disordered proteins or protein regions, prediction of intrinsic protein disorder from amino acid sequence has become an area of active research as witnessed in the 6th experiment on Critical Assessment of Techniques for Protein Structure Prediction (CASP6). Since the initial work by Romero et al. (Identifying disordered regions in proteins from amino acid sequences, IEEE Int. Conf. Neural Netw., 1997), our group has developed several predictors optimized for long disordered regions (>30 residues) with prediction accuracy exceeding 85%. However, these predictors are less successful on short disordered regions (≤30 residues). A probable cause is a length-dependent amino acid compositions and sequence properties of disordered regions. Results We proposed two new predictor models, VSL2-M1 and VSL2-M2, to address this length-dependency problem in prediction of intrinsic protein disorder. These two predictors are similar to the original VSL1 predictor used in the CASP6 experiment. In both models, two specialized predictors were first built and optimized for short (≤30 residues) and long disordered regions (>30 residues), respectively. A meta predictor was then trained to integrate the specialized predictors into the final predictor model. As the 10-fold cross-validation results showed, the VSL2 predictors achieved well-balanced prediction accuracies of 81% on both short and long disordered regions. Comparisons over the VSL2 training dataset via 10-fold cross-validation and a blind-test set of unrelated recent PDB chains indicated that VSL2 predictors were significantly more accurate than several existing predictors of intrinsic protein disorder. Conclusion The VSL2 predictors are applicable to disordered regions of any length and can accurately identify the short disordered regions that are often misclassified by our previous disorder predictors. The success of the VSL2 predictors further confirmed the previously observed differences in amino acid compositions and sequence properties between short and long disordered regions, and justified our approaches for modelling short and long disordered regions separately. The VSL2 predictors are freely accessible for non-commercial use at
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                Author and article information

                Contributors
                Role: ConceptualizationRole: InvestigationRole: MethodologyRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: InvestigationRole: MethodologyRole: Writing – original draft
                Role: InvestigationRole: MethodologyRole: Writing – review & editing
                Role: Data curationRole: MethodologyRole: Writing – review & editing
                Role: InvestigationRole: MethodologyRole: Writing – review & editing
                Role: InvestigationRole: Writing – review & editing
                Role: ConceptualizationRole: InvestigationRole: Writing – review & editing
                Role: InvestigationRole: MethodologyRole: Writing – review & editing
                Role: InvestigationRole: Writing – review & editing
                Role: Funding acquisitionRole: Project administrationRole: ResourcesRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS Genet
                PLoS Genet
                plos
                plosgen
                PLoS Genetics
                Public Library of Science (San Francisco, CA USA )
                1553-7390
                1553-7404
                6 November 2018
                November 2018
                : 14
                : 11
                : e1007696
                Affiliations
                [1 ] Institute for Developmental Biochemistry, University Medical Center, Göttingen, Germany
                [2 ] Laboratory of Metabolism, Department of Internal Medicine Specialties, Faculty of Medicine, University of Geneva, Switzerland
                [3 ] Institute of Human Genetics, University Medical Center, Göttingen, Germany
                Fred Hutchinson Cancer Research Center, UNITED STATES
                Author notes

                The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0001-7239-7404
                http://orcid.org/0000-0002-4621-4624
                http://orcid.org/0000-0002-1402-2628
                http://orcid.org/0000-0003-2433-1851
                http://orcid.org/0000-0001-9247-8586
                Article
                PGENETICS-D-18-00493
                10.1371/journal.pgen.1007696
                6219760
                30399145
                5b37e4a9-040e-4768-b090-0767f0d609a6
                © 2018 Krishnakumar et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 14 March 2018
                : 16 September 2018
                Page count
                Figures: 8, Tables: 0, Pages: 29
                Funding
                Funded by: German Egyptian Research Long-term Scholarship
                Award ID: 91664584
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001659, Deutsche Forschungsgemeinschaft;
                Award ID: DO 740/2-3
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001650, German Academic Exchange Service New Delhi;
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100001650, German Academic Exchange Service New Delhi;
                Award Recipient :
                This work was supported by a GZMB stipend, a GGNB bridging fund (SR), the German Academic Exchange Service, DAAD https://www.daad.de/en/) (PK, RP, HK), the Deutsche Forschungsgemeinschaft (DO 740/2-3) ( http://www.dfg.de) the GGNB Junior Group Stipend and the ‘‘Forschungsförderungsprogramm’’ of the University Medical Center Göttingen (RD). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
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