Proteins In Various Species Research Articles

Glycogen synthase activity in Candida albicans is partly controlled by the functional ortholog of Saccharomyces cerevisiae Gac1p.

To adapt to various host microenvironments, the human fungal pathogen Candida albicans possesses the capacity to accumulate and store glycogen as an internal carbohydrate source. In the model yeast Saccharomyces cerevisiae, ScGlc7p and ScGac1p are the serine/threonine type 1 protein phosphatase catalytic and regulatory subunits that control glycogen synthesis by altering the phosphorylation state of the glycogen synthase Gsy2p. Despite recent delineation of the glycogen synthesis pathway in C. albicans, the molecular events driving synthase activation are currently undefined. In this study, using a combination of microbiologic and genetic techniques, we determined that the protein encoded by uncharacterized gene C1_01140C, and not the currently annotated C. albicans Gac1p, is the major regulatory subunit involved in glycogen synthesis. C1_01140Cp contains a conserved GVNK motif observed across multiple starch/glycogen-binding proteins in various species, and alanine substitution of each residue in this motif significantly impaired glycogen accumulation in C. albicans. Fluorescent protein tagging and microscopy indicated that C1_01140Cp-GFPy colocalized with CaGlc7p-tdTomato and CaGsy1p-tdTomato accordingly. Co-immunoprecipitation assays further confirmed that C1_01140Cp associates with CaGlc7p and CaGsy1p during glycogen synthesis. Lastly, c1_01140cΔ/Δ exhibited colonization defects in a murine model of vulvovaginal candidiasis. Collectively, our data indicate that uncharacterized C1_01140Cp is the functional ortholog of the PPP1R subunit ScGac1p in C. albicans.IMPORTANCEThe capacity to synthesize glycogen offers microbes metabolic flexibility, including the fungal pathogen Candida albicans. In Saccharomyces cerevisiae, dephosphorylation of glycogen synthase by the ScGlc7p-containing phosphatase is a critical rate-limiting step in glycogen synthesis. Subunits, including ScGac1p, target ScGlc7p to α-1,4-glucosyl primers for efficient ScGsy2p synthase activation. However, this process in C. albicans had not been delineated. Here, we show that the C. albicans genome encodes for two homologous phosphatase-binding subunits, annotated CaGac1p and uncharacterized C1_01140Cp, both containing a GVNK motif required for polysaccharide affinity. Surprisingly, loss of CaGac1p only moderately reduced glycogen accumulation, whereas loss of C1_01140Cp ablated it. Fluorescence microscopy and co-immunoprecipitation approaches revealed that C1_01140Cp associates with CaGlc7p and CaGsy1p during glycogen synthesis. Moreover, C1_01140Cp contributed to fungal fitness at the vaginal mucosa during murine vaginitis. Therefore, this work demonstrates that glycogen synthase regulation is conserved in C. albicans and C1_01140Cp is the functional ortholog of ScGac1p.

Autophagy-related protein PlAtg3 participates in vegetative growth, sporangial cleavage, autophagy and pathogenicity of Peronophythora litchii

Litchi downy blight, caused by the plant pathogenic oomycete Peronophythora litchii, is one of the most devastating diseases on litchi and resulted in huge economic losses. Autophagy plays an essential role in the development and pathogenicity of the filamentous fungi. However, the function of autophagy in oomycetes remain elusive. Here, an autophagy-related protein Atg3 homolog PlAtg3 was identified and characterized in P. litchii. The absence of PlATG3 through the CRISPR/Cas9 gene replacement strategy compromised vegetative growth and sexual/asexual development. Cytological analyses revealed that the deletion of PlATG3 impaired autophagosome formation with monodansylcadaverine (MDC) staining and significantly disrupted zoospore release due to defects of sporangial cleavage with FM4-64 staining. Atg8 is considered to be an autophagy marker protein in various species. Western blot analysis indicated that PlAtg3 is involved in degradation of PlAtg8-PE. Interestingly, PlAtg3 was unable to interact with PlAtg8 in yeast two hybrid (Y2H) assays, possibly due to the absence of the Atg8 family interacting motif (AIM) in PlAtg3. Furthermore, pathogenicity assays revealed that the deletion of PlATG3 considerably reduced the virulence of P. litchii. Taken together, our data reveal that PlAtg3 plays an important role in radial growth, asexual/sexual development, sporangial cleavage and zoospore release, autophagosome formation, and pathogenicity in P. litchii. This study contributes to a better understanding of the pathogenicity mechanisms of P. litchii and provides insights for the development of more effective strategies to control oomycete diseases.

Synapse Formation and Function Across Species: Ancient Roles for CCP, CUB, and TSP-1 Structural Domains.

The appearance of synapses was a crucial step in the creation of the variety of nervous systems that are found in the animal kingdom. With increased complexity of the organisms came a greater number of synaptic proteins. In this review we describe synaptic proteins that contain the structural domains CUB, CCP, or TSP-1. These domains are found in invertebrates and vertebrates, and CUB and CCP domains were initially described in proteins belonging to the complement system of innate immunity. Interestingly, they are found in synapses of the nematode C. elegans, which does not have a complement system, suggesting an ancient function. Comparison of the roles of CUB-, CCP-, and TSP-1 containing synaptic proteins in various species shows that in more complex nervous systems, these structural domains are combined with other domains and that there is partial conservation of their function. These three domains are thus basic building blocks of the synaptic architecture. Further studies of structural domains characteristic of synaptic proteins in invertebrates such as C. elegans and comparison of their role in mammals will help identify other conserved synaptic molecular building blocks. Furthermore, this type of functional comparison across species will also identify structural domains added during evolution in correlation with increased complexity, shedding light on mechanisms underlying cognition and brain diseases.

Complex Networks of Prion-Like Proteins Reveal Cross Talk Between Stress and Memory Pathways in Plants.

Prions are often considered as molecular memory devices, generating reproducible memory of a conformational change. Prion-like proteins (PrLPs) have been widely demonstrated to be present in plants, but their role in plant stress and memory remains unexplored. In this work, we report the widespread presence of PrLPs in plants through a comprehensive meta-analysis of 39 genomes representing major taxonomic groups. We find diverse functional roles associated with these proteins in various species and term the full complement of PrLPs in a genome as its “prionome.” In particular, we found the rice prionome being significantly enriched in transposons/retrotransposons (Ts/RTRs) and identified over 60 rice PrLPs that were differentially regulated in stress and developmental responses. This prompted us to explore whether and to what extent PrLPs may build stress memory. By integrating the available rice interactome, transcriptome, and regulome data sets, we could find links between stress and memory pathways that would not have otherwise been discernible. Regulatory inferences derived from the superimposition of these data sets revealed a complex network and cross talk between PrLPs, transcription factors (TFs), and the genes involved in stress priming. This integrative meta-analysis connects transient and transgenerational memory mechanisms in plants with PrLPs, suggesting that plant memory may rely upon protein-based signals in addition to chromatin-based epigenetic signals. Taken together, our work provides important insights into the anticipated role of prion-like candidates in stress and memory, paving the way for more focused studies for validating the role of the identified PrLPs in memory acclimation.

Whey allergens: Influence of nonthermal processing treatments and their detection methods.

Whey and its components are recognized as value-added ingredients in infant formulas, beverages, sports nutritious foods, and other food products. Whey offers opportunities for the food industrial sector to develop functional foods with potential health benefits due to its unique physiological and functional attributes. Despite all the above importance, the consumption of whey protein (WP) can trigger hypersensitive reactions and is a constant threat for sensitive individuals. Although avoiding such food products is the most successful approach, there is still a chance of incorrect labeling and cross-contamination during food processing. As whey allergens in food products are cross-reactive, the phenomenon of homologous milk proteins of various species may escalate to a more serious problem. In this review, nonthermal processing technologies used to prevent and eliminate WP allergies are presented and discussed in detail. These processing technologies can either enhance or mitigate the impact of potential allergenicity. Therefore, the development of highly precise analytical technologies to detect and quantify the existence of whey allergens is of considerable importance. The present review is an attempt to cover all the updated approaches used for the detection of whey allergens in processed food products. Immunological and DNA-based assays are generally used for detecting allergenic proteins in processed food products. In addition, mass spectrometry is also employed as a preliminary technique for detection. We also highlighted the latest improvements in allergen detection toward biosensing strategies particularly immunosensors and aptasensors.

FSH1 encodes lysophospholipase activity in Saccharomyces cerevisiae.

To elucidate the role of FSH1 (family of serine hydrolase) in lipid homeostasis. Proteins in various species containing alpha/beta hydrolase domain are known to be involved in lipid metabolism. In silico analysis of the FSH1 gene in Saccharomyces cerevisiae revealed the presence of alpha/beta hydrolase domain (ABHD) and a lipase motif (GXSXG), however its function in lipid metabolism remained elusive. The overexpression of FSH1 in WT and fsh1Δ cells showed a significant reduction in the cellular phospholipid levels and an increase in the triacylglycerol levels and lipid droplet (LD) number. Furthermore, the purified recombinant protein Fsh1p was identified as a lysophospholipase that specifically acts on lysophosphatidylserine (LPS) and impacts the lipid homeostasis in S. cerevisiae. These results depicted that Fsh1p has a role on lipid homeostasis and is a lysophospholipase that hydrolyzes lysophosphatidylserine (LPS).

Deleted in azoospermia-associated protein 2 regulates innate immunity by stimulating Hippo signaling in crab

The Hippo-signaling pathway plays a critical role in both normal animal physiology and pathogenesis. Because pharmacological interventions targeting this pathway have diverse clinical implications, a better understanding of its regulation in various conditions and organisms is crucial. Here, we identified deleted in azoospermia-associated protein 2 (DAZAP2) in the Chinese mitten crab (Eriocheir sinensis), designated EsDAZAP2, as a Hippo-regulatory protein highly similar to proteins in various species of insects, fish, and mammals. We found that a bacterial infection significantly induces EsDAZAP2 expression, and an EsDAZAP2 knockdown both suppresses antimicrobial peptide (AMP) expression in vitro and results in increased viable bacterial counts and mortality in vivo, suggesting that EsDAZAP2 plays a critical role in innate immunity. Using yeast two-hybrid screening and co-immunoprecipitation assays, we found that EsDAZAP2 regulates the Toll pathway rather than the immune deficiency and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathways. Our findings also demonstrate that EsDAZAP2 binds to the Hippo protein, Salvador (Sav). Moreover, by examining the regulation of Dorsal, a transcription factor that regulates AMP expression in E. sinensis, we provide experimental evidence indicating that EsDAZAP2 promotes Hippo pathway activation in innate immunity, with EsDAZAP2 and Hippo binding to different Sav domains. To the best of our knowledge, this is the first report of a DAZAP2-regulated Hippo-signaling pathway operating in animal innate immunity.

Analysis of MYOC gene variants among sporadic patients with primary open-angle glaucoma

To screen for MYOC gene variants among sporadic patients with primary open angle glaucoma (POAG). For 398 patients with POAG, Sanger sequencing was applied to detect potential variants of the MYOC gene. Eight patients (2.0%) were found to harbor variations of the MYOC gene. These included five types of variants, among which c.667C>T (p.Pro223Ser) and c.1138G>T (p.Asp380Tyr) were novel. c.382C>T (p.Arg128Trp), c.1109C>T(p.Pro370Leu) and c.1130C>A (p.Thr377Lys) were previously associated with POAG. Alignment of amino acid sequences of MYOC proteins of various species revealed that the two novel variants have occurred at highly conserved positions. c.1138G>T was predicted to be possible pathogenic by Bioinformatic analysis. Two novel variants of the MYOC gene were detected among sporadic POAG patients, which enriched its variant spectrum.

Postsynaptic density proteins and their involvement in neurodevelopmental disorders.

Synaptic connections are essential for neural circuits in order to convey brain functions. The postsynaptic density (PSD) is a huge protein complex associated with postsynaptic membranes of excitatory synapses. In mammals, the PSD is composed of more than 1,000 proteins including receptors, scaffold proteins, signalling enzymes and cytoskeletal proteins. PSD proteins are crucial for synaptic transmission and plasticity. Proteomic studies have revealed the composition of PSD proteins in various species, brain regions and specific physiological conditions. Abnormalities with PSD proteins are linked to various neuropsychiatric diseases including neurodevelopmental disorders such as autism spectrum disorder and schizophrenia. Here, we review different kinds of proteomic studies of the PSD and the involvement of PSD proteins in physiological and pathological conditions.

Local Palmitoylation Cycles andSpecialized Membrane Domain Organization.

Palmitoylation is an evolutionally conserved lipid modification of proteins. Dynamic and reversible palmitoylation controls a wide range of molecular and cellular properties of proteins including the protein trafficking, protein function, protein stability, and specialized membrane domain organization. However, technical difficulties in (1) detection of palmitoylated substrate proteins and (2) purification and enzymology of palmitoylating enzymes have prevented the progress in palmitoylation research, compared with that in phosphorylation research. The recent development of proteomic and chemical biology techniques has unexpectedly expanded the known complement of palmitoylated proteins in various species and tissues/cells, and revealed the unique occurrence of palmitoylated proteins in membrane-bound organelles and specific membrane compartments. Furthermore, identification and characterization of DHHC (Asp-His-His-Cys) palmitoylating enzyme-substrate pairs have contributed to elucidating the regulatory mechanisms and pathophysiological significance of protein palmitoylation. Here, we review the recent progress in protein palmitoylation at the molecular, cellular, and in vivo level and discuss how locally regulated palmitoylation machinery works for dynamic nanoscale organization of membrane domains.

Structure and function of placental exchange surfaces in goodeid fishes (Teleostei: Atheriniformes).

The species of the family Goodeidae have evolved reproductive strategies involving intraovarian gestation, early evacuation of nearly yolk-exhausted embryos from the ovigerous tissue into the ovarian cavity, placental matrotrophy during intraluminal gestation, and the birth of highly developed fry. The inner ovarian lining becomes hypervascularized during gestational periods and functions as the maternal component of the placental association. Embryotrophic liquid is secreted by the inner ovarian epithelium into the ovarian cavity. Comparative electrophoretic analyses of embryotrophe and maternal blood serum provide evidence for the transfer of maternal serum proteins into the embryotrophe. Trophotaeniae, proctodaeal processes of the embryos, provide a surface for nutrient absorption. Endocytic activity was demonstrated by ingestion of unspecific tracer proteins in various species. Moreover, the trophotaenial absorptive cells (TACs) in Ameca splendens ingest various proteins or random copolymers conjugated to colloidal gold as well as radioiodinated proteins in a way that satisfies the criteria of receptor-mediated endocytosis. Several aminopeptidases (APs) on the surface of TACs were identified as protein binding sites as evidenced by inhibition of binding and uptake of marker proteins in the presence of AP substrates or AP inhibitors. Morphological adaptations of the embryonic circulatory system pertaining to nutrient and gas exchange were characterized. The embryonic epidermis comprises two layers of squamous cells closely underlain by a dense capillary net. Efficient gas exchange is facilitated by a thin embryotrophe-blood barrier of both the embryonic skin and the intraovarian lining.

The Molecular Evolution of the Pif Family Proteins in Various Species of Mollusks

Various novel proteins have been identified from many kinds of mollusk shells. Although such matrix proteins are believed to play important roles in the calcium carbonate crystal formation of shells, no common proteins that interact with calcium carbonate or that are involved in the molecular mechanisms behind shell formation have been identified. Pif consists of two proteins, Pif 80 and Pif 97, which are encoded by a single mRNA. Pif 80 was identified as a key acidic protein that regulates the formation of the nacreous layer in Pinctada fucata, while Pif 97 has von Willebrand factor type A (VWA) and chitin-binding domains. In this study, we identified Pif homologues from Pinctada margaritifera, Pinctada maxima, Pteria penguin, Mytilus galloprovincialis, and in the genome database of Lottia gigantea in order to compare their primary protein sequences. The VWA and chitin-binding domains are conserved in all Pif 97 homologues, whereas the amino acid sequences of the Pif 80 regions differ markedly among the species. Sequence alignment revealed the presence of a novel significantly conserved sequence between the chitin-binding domain and the C-terminus of Pif 97. Further examination of the Pif 80 regions suggested that they share a sequence that is similar to the laminin G domain. These results indicate that all Pif molecules in bivalves and gastropods may be derived from a common ancestral gene. These comparisons may shed light on the correlation between molecular evolution and morphology in mollusk shell microstructure.

Brassinosteroids and Auxin Down-Regulate DELLA Genes in Fiber Initiation and Elongation of Cotton

Plant hormones play important roles in cotton fiber growth and development. However, the interaction of phytohormones is largely unknown in fiber cells up to now. DELLA proteins are critical component in GA (gibberellic acid) signal transduction, which are also regulated by other phytohormones, such as auxin and ethylene. To understand the regulation of DELLA genes in cotton fiber growth and development, we cloned four DELLA genes from upland cotton fibers (Gossypium hirsutum L.), named GhGAI1, GhGAI2, GhGAI3, and GhGAI4. Alignment of the four predicted proteins with other reported DELLA proteins in various species displayed that they shared conserved domains and high homology. Expression profiles of the four GhGAIs in various tissues and organs as well as cotton fibers in different stages displayed that GhGAI1 has higher transcriptional levels than other GhGAIs in all detected samples. Furthermore, the expression level of GhGAI1 was significantly reduced in 0 dpa (day post anthesis) ovules by addition of IAA and epi-BL, and exogenous epi-BL decreased GhGAI1 level in 7 dpa fiber. Similarly, the levels of the other three GhGAIs in 0 dpa ovules and 7 dpa fibers were also regulated by applied phytohormones. In addition, the levels of GhGAI1 were higher in Xuzhou142 fl mutant (fuzzless-lintless) than in FL (Gossypium hirsutum vs. Xuzhou 142) from −1 to 3 dpa ovules, suggesting that GhGAI1 engaged in cotton fiber cell initiation. These results indicated that DELLA genes are involved in the process of fiber cell initiation and elongation regulated by different phytohormones.

Characteristics and evolution of the PUF gene family in Bombyx mori and 27 other species

The Pumilio protein is the founding member of the PUF family of RNA-binding proteins, which contains 8 repeat Puf domains and plays important roles during embryogenesis and post-embryogenesis by binding the Nanos response element (NRE) of specific target genes in eukaryotes. In addition, many other proteins containing the Puf domain were identified but with different functions from the Pumilio protein in various species. Taking advantage of the newly assembled genome sequences, in this study we performed a genome-wide analysis of PUF genes in silkworm and other 27 species. In the silkworm, three PUF genes were identified, named Bmpumilio, Bmpenguin and Bmnop by homology analysis. In fungi and animals, four evolutionarily conservational PUF gene families were identified, Group-A, -B, -C and -D. While Group-A, -C, and -D are present in all fungi and animals, Group-B was only identified in fungi. Interestingly, the number and features of the Puf domains are distinct in each group, suggesting different roles for these proteins in every group. The EST and microarray data showed that the mRNA of the three PUF genes can be widely detected in all tissues of the silkworm. Our results provide some new insights into the functions and evolutionary characteristics of PUF proteins.

Septic encephalopathy: Inflammation in man and mouse

Septic encephalopathy is a frequent complication of the sepsis syndrome, with no therapies available that can prevent the associated neurological dysfunction in humans. It is caused by a number of processes and networks going awry, the exact cellular and molecular mechanisms of which remain an enigma. Several mediators of inflammation have been assigned a key role in sepsis, including cytokines, chemokines and complement cascade. With the observations that brain dysfunction in a sepsis setting can be alleviated by regulation of the cytokines and complement proteins in various species of animals, optimism is building for a possible therapy of sepsis-damaged brain. This article reviewed the advances in the understanding of the underlying mechanisms causing pathology in SE, with an emphasis on the inflammatory and excitatory mediators such as the cytokines, complement proteins and neurotransmitters, investigating their potential as possible therapeutic targets.

UCHL-1 protein expression specifically marks spermatogonia in wild bovid testis

Spermatogonia are germ cells that initiate spermatogenesis in mammalian testis and they are the only cells in adult body capable of dividing both mitotically and meiotically. Therefore, isolation and preservation of spermatogonia can provide an alternative method for preservation of genetic pool of endangered animals. To achieve this objective, it is essential to identify markers that can specifically distinguish spermatogonia from other cells in the testis. In the present study, anti-ubiquitin C-terminal hydroxylase-1 (UCHL-1/PGP 9.5) antibody specifically recognized spermatogonia in the testis of wild and domestic bovid. The size of the UCHL-1 protein in various species of bovid testes was identical, and similar to that in mice. Furthermore, UCHL-1 staining could be utilized for the identification of spermatogonia in isolated testicular cells from wild bovids suggesting that UCHL-1 protein expression could be used as a specific marker for spermatogonia in bovid family.

Functional involvement of Tudor and dPRMT5 in the piRNA processing pathway in Drosophila germlines

In Drosophila, the PIWI proteins, Aubergine (Aub), AGO3, and Piwi are expressed in germlines and function in silencing transposons by associating with PIWI-interacting RNAs (piRNAs). Recent studies show that PIWI proteins contain symmetric dimethyl-arginines (sDMAs) and that dPRMT5/Capsuleen/DART5 is the modifying enzyme. Here, we show that Tudor (Tud), one of Tud domain-containing proteins, associates with Aub and AGO3, specifically through their sDMA modifications and that these three proteins form heteromeric complexes. piRNA precursor-like molecules are detected in these complexes. The expression levels of Aub and AGO3, along with their degree of sDMA modification, were not changed by tud mutations. However, the population of transposon-derived piRNAs associated with Aub and AGO3 was altered by tud mutations, whereas the total amounts of small RNAs on Aub and AGO3 was increased. Loss of dprmt5 did not change the stability of Aub, but impaired its association with Tud and lowered piRNA association with Aub. Thus, in germline cells, piRNAs are quality-controlled by dPRMT5 that modifies PIWI proteins, in tight association with Tud.

Crystal structure of ST2348, a CBS domain protein, from hyperthermophilic archaeon Sulfolobus tokodaii

The crystal structure of a hypothetical protein ST2348 (GI: 47118305) from the hyperthermophilic bacteria Sulfolobus tokodaii has been determined using X-ray crystallography. The protein consists of two CBS (cystathione β synthase) domains, whose function has been analyzed and reported here. PSI-BLAST shows a conservation of this domain in about 100 proteins in various species. However, none of the close homologs of ST2348 have been functionally characterized so far. Structure and sequence comparison of ST2348 with human AMP-kinase γ1 subunit and the CBS domain pair of bacterial IMP dehydrogenase is suggestive of its binding to AMP and ATP. A highly conserved residue Asp118, located in a negatively charged patch near the ligand binding cleft, could serve as a site for phosphorylation similar to that found in the chemotatic signal protein CheY and thereby ST2348 can function as a signal transduction molecule.

Apolipoprotein E phylogeny and evolution

Among apolipoproteins, apolipoprotein E (Apo E) plays a pivotal role in lipid transport and is proposed to be involved in neural repair. Because of a long divergence history of apolipoproteins, it is unclear how Apo E evolved in time. To investigate relationships among Apo E proteins, we used the information from molecular data and analysed the phylogeny of Apo E proteins from various species. Several phylogenetic trees were generated by using both character-based and distance-based phylogenetic methods. Apo E sequences of fish and frog were found to be less related to the Apo E sequences of other species. The most likely ancestor of Apo E among 18 organisms was estimated to be the Apo E of frog. Members of the groups formed by the Apo E proteins of various species shared similar feeding habits and diet. It may be suggested that Apo E evolution and very likely the evolution of other apolipoproteins are influenced by the organism's feeding environment and diet.

A β-Hairpin Comprising the Nuclear Localization Sequence Sustains the Self-associated States of Nucleosome Assembly Protein 1

The histone chaperone nucleosome assembly protein 1 (NAP1) is implicated in histone shuttling as well as nucleosome assembly and disassembly. Under physiological conditions, NAP1 dimers exist in a mixture of various high-molecular-weight oligomers whose size may be regulated by the cell cycle-dependent concentration of NAP1. Both the functional and structural significance of the observed oligomers are unknown. We have resolved the molecular mechanism by which yeast NAP1 (yNAP1) dimers oligomerize by applying x-ray crystallographic, hydrodynamic, and functional approaches. We found that an extended β-hairpin that protrudes from the compact core of the yNAP1 dimer forms a stable β-sheet with β-hairpins of neighboring yNAP1 dimers. Disruption of the β-hairpin (whose sequence is conserved among NAP1 proteins in various species) by the replacement of one or more amino acids with proline results in complete loss of yNAP1 dimer oligomerization. The in vitro functions of yNAP1 remain unaffected by the mutations. We have thus identified a conserved structural feature of NAP1 whose function, in addition to presenting the nuclear localization sequence, appears to be the formation of higher-order oligomers.