ACI-35 AND AADVAC1 ACTIVE IMMUNOTHERAPY AS PREVENTATIVE TREATMENT OPTIONS FOR CHRONIC TRAUMATIC ENCEPHALOPATHY

One of the most common, as well as one of the most dangerous injuries amongst athletes today is mild traumatic brain injury (mTBI), commonly known as concussion. Aside from physical symptoms such as nausea, dizziness, and headaches; concussions have can have longterm effects on brain physiology. A common neurological disease that can result from multiple concussions is Chronic Traumatic Encephalopathy (CTE), characterized by symptoms such as severe depression, anxiety, confusion, and aggression; amongst others.1 On the cellular level, CTE is classified by a unique pathway that leads to the hyperphosphorylation of tau protein and subsequent clumping of tau-containing neurofibrillary tangles (NFTs).2 Tau hyperphosphorylation and aggregation is also a key characteristic of a similar neurological condition, Alzheimer’s Disease (AD). Emerging active immunization treatment methods, including the ACI-35 and AADVac-1 vaccines, have been clinically tested in Alzheimer\u27s patients with the goal of reducing hyperphosphorylation of tau and its effects.3 Because of the similarity between the two diseases, this paper takes a look at the possibility of applying this active immunotherapy treatment to patients who are likely to develop CTE

Genomic, Biochemical, and Modeling Analyses of Asparagine Synthetases from Wheat

Asparagine synthetase activity in cereals has become an important issue with the discovery that free asparagine concentration determines the potential for formation of acrylamide, a probably carcinogenic processing contaminant, in baked cereal products. Asparagine synthetase catalyses the ATP-dependent transfer of the amino group of glutamine to a molecule of aspartate to generate glutamate and asparagine. Here, asparagine synthetase-encoding polymerase chain reaction (PCR) products were amplified from wheat (Triticum aestivum) cv. Spark cDNA. The encoded proteins were assigned the names TaASN1, TaASN2, and TaASN3 on the basis of comparisons with other wheat and cereal asparagine synthetases. Although very similar to each other they differed slightly in size, with molecular masses of 65.49, 65.06, and 66.24 kDa, respectively. Chromosomal positions and scaffold references were established for TaASN1, TaASN2, and TaASN3, and a fourth, more recently identified gene, TaASN4. TaASN1, TaASN2, and TaASN4 were all found to be single copy genes, located on chromosomes 5, 3, and 4, respectively, of each genome (A, B, and D), although variety Chinese Spring lacked a TaASN2 gene in the B genome. Two copies of TaASN3 were found on chromosome 1 of each genome, and these were given the names TaASN3.1 and TaASN3.2. The TaASN1, TaASN2, and TaASN3 PCR products were heterologously expressed in Escherichia coli (TaASN4 was not investigated in this part of the study). Western blot analysis identified two monoclonal antibodies that recognized the three proteins, but did not distinguish between them, despite being raised to epitopes SKKPRMIEVAAP and GGSNKPGVMNTV in the variable C-terminal regions of the proteins. The heterologously expressed TaASN1 and TaASN2 proteins were found to be active asparagine synthetases, producing asparagine and glutamate from glutamine and aspartate. The asparagine synthetase reaction was modeled using SNOOPY® software and information from the BRENDA database to generate differential equations to describe the reaction stages, based on mass action kinetics. Experimental data from the reactions catalyzed by TaASN1 and TaASN2 were entered into the model using Copasi, enabling values to be determined for kinetic parameters. Both the reaction data and the modeling showed that the enzymes continued to produce glutamate even when the synthesis of asparagine had ceased due to a lack of aspartate.Rothamsted Research by Shanghai Agriculture Applied Technology Development Program, China; Shanghai Academy of Agricultural Sciences, Shanghai, China; Biotechnology and Biological Sciences Research Council (BBSRC) of the United Kingdom; Rothamsted Research by the BBS

Enhancement of Heat Stability and Kinetic Parameters of the Maize Endosperm ADP-Glucose Pyrophosphorylase by Mutagenesis of Amino Acids in the Small Subunit With High B Factors

ADP-glucose pyrophosphorylase (AGPase) is an important enzyme in starch synthesis and previous studies showed that the heat lability of this enzyme is a determinant to starch synthesis in the maize endosperm and, in turn, seed yield. Here, amino acids in the AGPase endosperm small subunit with high B-factors were mutagenized and individual changes enhancing heat stability and/or kinetic parameters in an Escherichia coli expression system were chosen. Individual mutations were combined and analyzed. One triple mutant, here termed Bt2-BF, was chosen for further study. Combinations of this heat stable, 3-PGA-independent small subunit variant with large subunits also heat stable yielded complex patterns of heat stability and kinetic and allosteric properties. Interestingly, two of the three changes reside in a protein motif found only in AGPases that exhibit high sensitivity to 3-PGA. While not the 3-PGA binding site, amino acid substitutions in this region significantly alter 3-PGA activation kinetics

Functions of maize genes encoding pyruvate phosphate dikinase in developing endosperm

Maize opaque2 (o2) mutations are beneficial for endosperm nutritional quality but cause negative pleiotropic effects for reasons that are not fully understood. Direct targets of the bZIP transcriptional regulator encoded by o2 include pdk1 and pdk2 that specify pyruvate phosphate dikinase (PPDK). This enzyme reversibly converts AMP, pyrophosphate, and phosphoenolpyruvate to ATP, orthophosphate, and pyruvate and provides diverse functions in plants. This study addressed PPDK function in maize starchy endosperm where it is highly abundant during grain fill. pdk1 and pdk2 were inactivated individually by transposon insertions, and both genes were simultaneously targeted by endosperm-specific RNAi. pdk2 accounts for the large majority of endosperm PPDK, whereas pdk1 specifies the abundant mesophyll form. The pdk1- mutation is seedling-lethal, indicating that C4 photosynthesis is essential in maize. RNAi expression in transgenic endosperm eliminated detectable PPDK protein and enzyme activity. Transgenic kernels weighed the same on average as nontransgenic siblings, with normal endosperm starch and total N contents, indicating that PPDK is not required for net storage compound synthesis. An opaque phenotype resulted from complete PPDK knockout, including loss of vitreous endosperm character similar to the phenotype conditioned by o2-. Concentrations of multiple glycolytic intermediates were elevated in transgenic endosperm, energy charge was altered, and starch granules were more numerous but smaller on average than normal. The data indicate that PPDK modulates endosperm metabolism, potentially through reversible adjustments to energy charge, and reveal that o2- mutations can affect the opaque phenotype through regulation of PPDK in addition to their previously demonstrated effects on storage protein gene expression

Mitochondrial Zea mays Brittle1-1 Is a Major Determinant of the Metabolic Fate of Incoming Sucrose and Mitochondrial Function in Developing Maize Endosperms

[EN] Zea mays Brittle1-1 (ZmBT1-1) is an essential component of the starch biosynthetic machinery in maize endosperms, enabling ADPglucose transport from cytosol to amyloplast in exchange for AMP or ADP. Although ZmBT1-1 has been long considered to be an amyloplast-specific marker, evidence has been provided that ZmBT1-1 is dually localized to plastids and mitochondria (Bahaji et al., 2011b). The mitochondrial localization of ZmBT1-1 suggested that this protein may have as-yet unidentified function(s). To understand the mitochondrial ZmBT1-1 function(s), we produced and characterized transgenic Zmbt1-1 plants expressing ZmBT1-1 delivered specifically to mitochondria. Metabolic and differential proteomic analyses showed down-regulation of sucrose synthase (SuSy)-mediated channeling of sucrose into starch metabolism, and up-regulation of the conversion of sucrose breakdown products generated by cell wall invertase (CWI) into ethanol and alanine, in Zmbt1-1 endosperms compared to wild-type. Electron microscopic analyses of Zmbt1-1 endosperm cells showed gross alterations in the mitochondrial ultrastructure. Notably, the protein expression pattern, metabolic profile, and aberrant mitochondrial ultrastructure of Zmbt1-1 endosperms were rescued by delivering ZmBT1-1 specifically to mitochondria. Results presented here provide evidence that the reduced starch content in Zmbt1-1 endosperms is at least partly due to (i) mitochondrial dysfunction, (ii) enhanced CWI-mediated channeling of sucrose into ethanol and alanine metabolism, and (iii) reduced SuSy-mediated channeling of sucrose into starch metabolism due to the lack of mitochondrial ZmBT1-1. Our results also strongly indicate that (a) mitochondrial ZmBT1-1 is an important determinant of the metabolic fate of sucrose entering the endosperm cells, and (b) plastidic ZmBT1-1 is not the sole ADPglucose transporter in maize endosperm amyloplasts. The possible involvement of mitochondrial ZmBT1-1 in exchange between intramitochondrial AMP and cytosolic ADP is discussed.This research was partially supported by the grants BIO2010-18239, BI2013-49125-C2-2-P and BIO2016-78747-P from the Comisión Interministerial de Ciencia y Tecnología and Fondo Europeo de Desarrollo Regional (Spain) and by the ERDF project Plants as a tool for sustainable global development (No. CZ.02.1.01/0.0/0.0/16_019/0000827).Bahaji, A.; Muñoz, FJ.; Seguí-Simarro, JM.; Camacho-Fernández, C.; Rivas-Sendra, A.; Parra Vega, V.; Ovecka, M.... (2019). Mitochondrial Zea mays Brittle1-1 Is a Major Determinant of the Metabolic Fate of Incoming Sucrose and Mitochondrial Function in Developing Maize Endosperms. 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PYRIMIDINE AND PURINE BIOSYNTHESIS AND DEGRADATION IN PLANTS. Annual Review of Plant Biology, 57(1), 805-836. doi:10.1146/annurev.arplant.57.032905.10542

Detection of antibodies directed at M. hyorhinis p37 in the serum of men with newly diagnosed prostate cancer

<p>Abstract</p> <p>Background</p> <p>Recent epidemiologic, genetic, and molecular studies suggest infection and inflammation initiate certain cancers, including cancers of the prostate. Over the past several years, our group has been studying how mycoplasmas could possibly initiate and propagate cancers of the prostate. Specifically, <it>Mycoplasma hyorhinis </it>encoded protein p37 was found to promote invasion of prostate cancer cells and cause changes in growth, morphology and gene expression of these cells to a more aggressive phenotype. Moreover, we found that chronic exposure of benign human prostate cells to <it>M. hyorhinis </it>resulted in significant phenotypic and karyotypic changes that ultimately resulted in the malignant transformation of the benign cells. In this study, we set out to investigate another potential link between mycoplasma and human prostate cancer.</p> <p>Methods</p> <p>We report the incidence of men with prostate cancer and benign prostatic hyperplasia (BPH) being seropositive for <it>M. hyorhinis</it>. Antibodies to <it>M. hyorhinis </it>were surveyed by a novel indirect enzyme-linked immunosorbent assay (ELISA) in serum samples collected from men presenting to an outpatient Urology clinic for BPH (N = 105) or prostate cancer (N = 114) from 2006-2009.</p> <p>Results</p> <p>A seropositive rate of 36% in men with BPH and 52% in men with prostate cancer was reported, thus leading us to speculate a possible connection between <it>M. hyorhinis </it>exposure with prostate cancer.</p> <p>Conclusions</p> <p>These results further support a potential exacerbating role for mycoplasma in the development of prostate cancer.</p

Leishmania infantum Asparagine Synthetase A Is Dispensable for Parasites Survival and Infectivity

A growing interest in asparagine (Asn) metabolism has currently been observed in cancer and infection fields. Asparagine synthetase (AS) is responsible for the conversion of aspartate into Asn in an ATP-dependent manner, using ammonia or glutamine as a nitrogen source. There are two structurally distinct AS: the strictly ammonia dependent, type A, and the type B, which preferably uses glutamine. Absent in humans and present in trypanosomatids, AS-A was worthy of exploring as a potential drug target candidate. Appealingly, it was reported that AS-A was essential in Leishmania donovani, making it a promising drug target. In the work herein we demonstrate that Leishmania infantum AS-A, similarly to Trypanosoma spp. and L. donovani, is able to use both ammonia and glutamine as nitrogen donors. Moreover, we have successfully generated LiASA null mutants by targeted gene replacement in L. infantum, and these parasites do not display any significant growth or infectivity defect. Indeed, a severe impairment of in vitro growth was only observed when null mutants were cultured in asparagine limiting conditions. Altogether our results demonstrate that despite being important under asparagine limitation, LiAS-A is not essential for parasite survival, growth or infectivity in normal in vitro and in vivo conditions. Therefore we exclude AS-A as a suitable drug target against L. infantum parasites