On the role of residue phosphorylation in 14-3-3 partners: AANAT as a case study

Twenty years ago, a novel concept in protein structural biology was discovered: the intrinsically disordered regions (IDRs). These regions remain largely unstructured under native conditions and the more are studied, more properties are attributed to them. Possibly, one of the most important is their ability to conform a new type of protein-protein interaction. Besides the classical domain-to-domain interactions, IDRs follow a ´fly-casting´ model including ´induced folding´. Unfortunately, it is only possible to experimentally explore initial and final states. However, the complete movie of conformational changes of protein regions and their characterization can be addressed by in silico experiments. Here, we simulate the binding of two proteins to describe how the phosphorylation of a single residue modulates the entire process. 14-3-3 protein family is considered a master regulator of phosphorylated proteins and from a modern point-of-view, protein phosphorylation is a three component system, with writers (kinases), erasers (phosphatases) and readers. This later biological role is attributed to the 14-3-3 protein family. Our molecular dynamics results show that phosphorylation of the key residue Thr31 in a partner of 14-3-3, the aralkylamine N-acetyltransferase, releases the fly-casting mechanism during binding. On the other hand, the non-phosphorylation of the same residue traps the proteins, systematically and repeatedly driving the simulations into wrong protein-protein conformations.Fil: Masone, Diego Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina. Universidad Nacional de Cuyo. Facultad de Ingeniería; ArgentinaFil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Exactas y Naturales; Argentin

Controllability of protein-protein interaction phosphorylation-based networks: Participation of the hub 14-3-3 protein family

Posttranslational regulation of protein function is an ubiquitous mechanism in eukaryotic cells. Here, we analyzed biological properties of nodes and edges of a human protein-protein interaction phosphorylation-based network, especially of those nodes critical for the network controllability. We found that the minimal number of critical nodes needed to control the whole network is 29%, which is considerably lower compared to other real networks. These critical nodes are more regulated by posttranslational modifications and contain more binding domains to these modifications than other kinds of nodes in the network, suggesting an intra-group fast regulation. Also, when we analyzed the edges characteristics that connect critical and non-critical nodes, we found that the former are enriched in domain-to-eukaryotic linear motif interactions, whereas the later are enriched in domain-domain interactions. Our findings suggest a possible structure for protein-protein interaction networks with a densely interconnected and self-regulated central core, composed of critical nodes with a high participation in the controllability of the full network, and less regulated peripheral nodes. Our study offers a deeper understanding of complex network control and bridges the controllability theorems for complex networks and biological protein-protein interaction phosphorylation-based networked systems.Fil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Flores, Gabriel. Eventioz/eventbrite Company; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Differential expression and accumulation of 14-3-3 paralogs in 3T3-L1 preadipocytes and differentiated cells

The 14-3-3 protein family interacts with more than 2000 different proteins in mammals, as a result of its specific phospho-serine/phospho-threonine binding activity. Seven paralogs are strictly conserved in mammalian species. Here, we show that during adipogenic differentiation of 3T3-L1 preadipocytes, the level of each 14-3-3 protein paralog is regulated independently. For instance 14-3-3β, γ, and η protein levels are increased compared to untreated cells. In contrast, 14-3-3ε protein levels decreased after differentiation while others remained constant. In silico analysis of the promoter region of each gene showed differences that explain the results obtained at mRNA and protein levels.Fil: Gojanovich, Aldana Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Plasma Biochemistries and Morphometric Indices of Body Condition in Imperial Cormorant (Phalacrocorax atriceps) Chicks

Plasma biochemistries provide a complementary method for assessing physiological and nutritional status of free-ranging wild birds. Triglycerides, total protein and alkaline phosphatase were determined in 110 free-living Imperial Cormorant (Phalacrocorax atriceps) chicks aged 16-35 days, at Punta León (Argentina) during 2010 and 2011. Body mass at 30 days of age ("pre-fledging body condition", 2010 only) and body mass corrected by tarsus length at the time of blood sampling ("current body condition", 2011 only) were also determined. Variability of parameters by sex, hatching order, survival, age and breeding season was assessed, and the relationship between biochemical and morphometric indices was also explored. Morphometric indices were higher in A-chicks (pre-fledging body condition also varied with sex), and explained 35-55% of B-chick survival. Biochemistries differed significantly between breeding seasons, being higher in 2011. Alkaline phosphatase increased with age, and total protein was higher in A-chicks. Triglycerides and total protein accounted for 26% and 30%, respectively, of variation in current body condition; however, they did not forecast pre-fledging body condition. Lastly, total protein levels predicted B-chick survival (higher levels in surviving B-chicks), but their prognostic value was relatively low. The results suggest that unlike morphometric indices, the biochemistries chosen are valuable to assess individual body condition at the time of sampling, yet their applicability for predicting chick survival requires further evaluation.Fil: Gallo, Luciana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; ArgentinaFil: Quintana, Flavio Roberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; ArgentinaFil: Svagelj, Walter Sergio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencia Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Uhart, Marcela María. University of California; Estados Unido

Protein intrinsic disorder and network connectivity. The case of 14-3-3 proteins

The understanding of networks is a common goal of an unprecedented array of traditional disciplines. One of the protein network properties most influenced by the structural contents of its nodes is the inter-connectivity. Recent studies in which structural information was included into the topological analysis of protein networks revealed that the content of intrinsic disorder in the nodes could modulate the network topology, rewire networks, and change their inter-connectivity, which is defined by its clustering coefficient. Here, we review the role of intrinsic disorder present in the partners of the highly conserved 14-3-3 protein family on its interaction networks. The 14-3-3s are phospho-serine/threonine binding proteins that have strong influence in the regulation of metabolism and signal transduction networks. Intrinsic disorder increases the clustering coefficients, namely the inter-connectivity of the nodes within each 14-3-3 paralog networks. We also review two new ideas to measure intrinsic disorder independently of the primary sequence of proteins, a thermodynamic model and a method that uses protein structures and their solvent environment. This new methods could be useful to explain unsolved questions about versatility and fixation of intrinsic disorder through evolution. The relation between the intrinsic disorder and network topologies could be an interesting model to investigate new implicitness of the graph theory into biology.Fil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas ; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas ; Argentin

Exposure to selected pathogens in Geoffroy's cats and domestic carnivores from central Argentina

Wild carnivores share a high percentage of parasites and viruses with closely related domestic carnivores. Because of increased overlap and potential contact with domestic species, we conducted a retrospective serosurvey for 11 common carnivore pathogens in 40 Geoffroy's cats (Leopardus geoffroyi) sampled between 2000 and 2008 within or near two protected areas in central Argentina (Lihué Calel National Park, La Pampa, and Campos del Tuyú National Park, Buenos Aires), as well as five domestic cats and 11 domestic dogs from cattle ranches adjacent to Lihué Calel Park. Geoffroy's cats had detectable antibody to canine distemper virus (CDV), feline calicivirus (FCV), feline coronavirus, feline panleukopenia virus (FPV), Toxoplasma gondii, Leptospira interrogans (serovars Ictero/Icter and Ballum), and Dirofilaria immitis. None of the wild cats had antibodies to feline herpesvirus, feline immunodeficiency virus (FIV), feline leukemia virus, or rabies virus. Domestic dogs had antibodies to CDV, canine adenovirus, canine herpesvirus, and canine parvovirus. Antibodies to FPV, FCV, FIV, and T. gondii were found in domestic cats.We provide the first data on exposure of free-ranging Geoffroy's cats to pathogens at two sites within the core area of the species distribution range, including the first report of antibodies to CDV in this species. We encourage continued monitoring for diseases in wild and domestic carnivores as well as preventive health care for domestic animals, particularly in park buffer zones where overlap is greatest.Fil: Uhart, Marcela María. ildlife Conservation Society; ArgentinaFil: Rago, María Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Ecología, Genética y Evolución de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Ecología, Genética y Evolución de Buenos Aires; ArgentinaFil: Marull, Carolina A.. ildlife Conservation Society; ArgentinaFil: Ferreyra, Hebe del Valle. ildlife Conservation Society; ArgentinaFil: Pereira, Javier Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Museo Argentino de Ciencias Naturales “Bernardino Rivadavia”; Argentin

14-3-3γ silencing impairs osteogenic differentiation of human adipose derived-mesenchymal stem cells

14-3-3proteins constitute a family of regulatory molecules that participatein a plethora of cellular processes mainly through protein-proteininteractions. Even though 14-3-3 protein family members show somefunctional redundancy, there is growing evidence that indicatesevolutionary and biochemical diversity. Consistent with theliterature, previous research from our laboratory showed thatexpression levels of 14-3-3 paralogs are independently regulatedduring the adipogenesis and osteogenesis of human adiposederived-mesenchymal stem cells (hASCs). In the current work, we useda validated approach to isolate hASCs and studied the implication of14-3-3γon the osteogenic commitment of these cells. To address this purpose,we delivered a 14-3-3γshRNA construct into hASCs by pAd-BLOCKiT, an adenoviral vectorcontaining a human U6 promoter, and examined the effect on thedifferentiation potential into osteoblasts. The latter was evaluatedby: i) measuring alkaline phosphatase (ALP) activity, an early-stageosteoblast differentiation biomarker, and ii) detectingRunt-related transcription factor 2 (Runx2, master regulator of boneformation) protein levels. Cells were either maintained for 14 dayswith standard growth media (control, low glucose DMEM; 5% FBS) orinduced with an osteogenic differentiation medium (ODM; an optimizeddrug cocktail that includes dexamethasone, β-glycerophosphate,and 2-phospho-L-ascorbic acid). Our results clearly showed a decreasein both Runx2 protein levels and ALP activity in 14-3-3γdepleted hASCs. This also accords with our earlier observations,which showed that reduced expression of 14-3-3γhad a negative impact on the osteoblastic transdifferentiation ofNIH3T3-L1 cells. Taken together, these findings suggest a regulatoryrole for 14-3-3γin hASC differentiation to the osteogenic lineage.Fil: Rivera, Lautaro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaLVII Reunión Anual de la Sociedad Argentina de Investigaciónes en Bioquímica y Biología MolecularMendozaArgentinaSociedad Argentina de Investigaciónes en Bioquímica y Biología Molecula

Freely Available Tool (FAT) for automated quantification of lipid droplets in stained cells

In this study, wepropose an automatic procedure for digital image processing. Wedescribe a method that can efficiently quantify and characterizelipid droplets distributions in different cell types in culture.Prospectively, the lipid droplets detection method described in thiswork could be applied to static or time-lapse data, collected with asimple visible light or fluorescence microscopy equipment. Fullyautomated algorithms were implemented in Octave, a freely availablescientific package.Fil: Masone, Diego Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentina. Universidad Nacional de Cuyo. Facultad de Ingeniería; ArgentinaFil: Gojanovich, Aldana Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Frontini López, Yesica Romina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: del Veliz, Samanta. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Uhart, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Bustos, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; Argentin

Understanding Free-Roaming Horse Conflicts with Wildlife in the Great Basin

Horses and burros were introduced to North America in the 1600s with European missionaries and explorers. Over time, abandoned and released horses and burros formed herds and by the time European-American settlers began to explore North America in the 1700s, these free-roaming horses and burros had adapted to their habitat and been incorporated into Native American culture. By the mid-1900s, most Americans considered these horses as wild and symbols of freedom and beauty. In 1971, Congress passed the Wild and Free-Roaming Horses and Burros Act (Public Law 73-482) to provide federal protection for free-roaming horses and burros (also known as wild horses and burros) in the western United States. In 1978, the Act was amended (Public Law 95-514) to require the Bureau of Land Management (BLM) to “determine appropriate management levels (AMLs) for wild horses and burros on [designated] public lands.” The Bureau of Land Management set AML at 26,715 wild horses and burros on 29 million acres of public land across 10 western states. The U.S. Forest Service was also tasked with managing over 7,100 wild horses and 900 burros on 53 wild horse territories (USFS, 2020). Managing free-roaming horses and burros on public lands has its challenges. In this article, we explain some of the potential conflicts free-roaming horses create with native wildlife on western public lands