Aminolevulinate synthase 2 mediates erythrocyte differentiation by regulating larval globin expression during Xenopus primary hematopoiesis

AbstractHemoglobin synthesis by erythrocytes continues throughout a vertebrate’s lifetime. The mechanism of mammalian heme synthesis has been studied for many years; aminolevulinate synthase 2 (ALAS2), a heme synthetase, is associated with X-linked dominant protoporphyria in humans. Amphibian and mammalian blood cells differ, but little is known about amphibian embryonic hemoglobin synthesis. We investigated the function of the Xenopus alas2 gene (Xalas2) in primitive amphibian erythrocytes and found that it is first expressed in primitive erythroid cells before hemoglobin alpha 3 subunit (hba3) during primary hematopoiesis and in the posterior ventral blood islands at the tailbud stage. Xalas2 is not expressed during secondary hematopoiesis in the dorsal lateral plate. Hemoglobin was barely detectable by o-dianisidine staining and hba3 transcript levels decreased in Xalas2-knockdown embryos. These results suggest that Xalas2 might be able to synthesize hemoglobin during hematopoiesis and mediate erythrocyte differentiation by regulating hba3 expression in Xenopus laevis

アフリカツメガエル一次造血期の赤芽球分化を制御するXalas2遺伝子の役割

筑波大学 (University of Tsukuba)201

Loss of ALS2/Alsin Exacerbates Motor Dysfunction in a SOD1H46R-Expressing Mouse ALS Model by Disturbing Endolysosomal Trafficking

BACKGROUND: ALS2/alsin is a guanine nucleotide exchange factor for the small GTPase Rab5 and involved in macropinocytosis-associated endosome fusion and trafficking, and neurite outgrowth. ALS2 deficiency accounts for a number of juvenile recessive motor neuron diseases (MNDs). Recently, it has been shown that ALS2 plays a role in neuroprotection against MND-associated pathological insults, such as toxicity induced by mutant Cu/Zn superoxide dismutase (SOD1). However, molecular mechanisms underlying the relationship between ALS2-associated cellular function and its neuroprotective role remain unclear. METHODOLOGY/PRINCIPAL FINDINGS: To address this issue, we investigated the molecular and pathological basis for the phenotypic modification of mutant SOD1-expressing mice by ALS2 loss. Genetic ablation of Als2 in SOD1(H46R), but not SOD1(G93A), transgenic mice aggravated the mutant SOD1-associated disease symptoms such as body weight loss and motor dysfunction, leading to the earlier death. Light and electron microscopic examinations revealed the presence of degenerating and/or swollen spinal axons accumulating granular aggregates and autophagosome-like vesicles in early- and even pre-symptomatic SOD1(H46R) mice. Further, enhanced accumulation of insoluble high molecular weight SOD1, poly-ubiquitinated proteins, and macroautophagy-associated proteins such as polyubiquitin-binding protein p62/SQSTM1 and a lipidated form of light chain 3 (LC3-II), emerged in ALS2-deficient SOD1(H46R) mice. Intriguingly, ALS2 was colocalized with LC3 and p62, and partly with SOD1 on autophagosome/endosome hybrid compartments, and loss of ALS2 significantly lowered the lysosome-dependent clearance of LC3 and p62 in cultured cells. CONCLUSIONS/SIGNIFICANCE: Based on these observations, although molecular basis for the distinctive susceptibilities to ALS2 loss in different mutant SOD1-expressing ALS models is still elusive, disturbance of the endolysosomal system by ALS2 loss may exacerbate the SOD1(H46R)-mediated neurotoxicity by accelerating the accumulation of immature vesicles and misfolded proteins in the spinal cord. We propose that ALS2 is implicated in endolysosomal trafficking through the fusion between endosomes and autophagosomes, thereby regulating endolysosomal protein degradation in vivo

Different Human Copper-Zinc Superoxide Dismutase Mutants, SOD1G93A and SOD1H46R, Exert Distinct Harmful Effects on Gross Phenotype in Mice

Amyotrophic lateral sclerosis (ALS) is a heterogeneous group of fatal neurodegenerative diseases characterized by a selective loss of motor neurons in the brain and spinal cord. Creation of transgenic mice expressing mutant Cu/Zn superoxide dismutase (SOD1), as ALS models, has made an enormous impact on progress of the ALS studies. Recently, it has been recognized that genetic background and gender affect many physiological and pathological phenotypes. However, no systematic studies focusing on such effects using ALS models other than SOD1G93A mice have been conducted. To clarify the effects of genetic background and gender on gross phenotypes among different ALS models, we here conducted a comparative analysis of growth curves and lifespans using congenic lines of SOD1G93A and SOD1H46R mice on two different genetic backgrounds; C57BL/6N (B6) and FVB/N (FVB). Copy number of the transgene and their expression between SOD1G93A and SOD1H46R lines were comparable. B6 congenic mutant SOD1 transgenic lines irrespective of their mutation and gender differences lived longer than corresponding FVB lines. Notably, the G93A mutation caused severer disease phenotypes than did the H46R mutation, where SOD1G93A mice, particularly on a FVB background, showed more extensive body weight loss and earlier death. Gender effect on survival also solely emerged in FVB congenic SOD1G93A mice. Conversely, consistent with our previous study using B6 lines, lack of Als2, a murine homolog for the recessive juvenile ALS causative gene, in FVB congenic SOD1H46R, but not SOD1G93A, mice resulted in an earlier death, implying a genetic background-independent but mutation-dependent phenotypic modification. These results indicate that SOD1G93A- and SOD1H46R-mediated toxicity and their associated pathogenic pathways are not identical. Further, distinctive injurious effects resulted from different SOD1 mutations, which are associated with genetic background and/or gender, suggests the presence of several genetic modifiers of disease expression in the mouse genome

Pressure Loss Characteristics in Rotating Coolant Passages of Gas Turbine (Effect of Inlet Flow Angle in Intermediate Shaft)

ABSTRACT Pressure loss characteristics of flow inside of gas turbine intermediate shaft have been investigated experimentally using a rotating test facility with disks having diameter of 0.5m and maximum rotational speed of 4800rpm. With controlling rotational Reynolds number and coolant flow rate (Rotation number), three types of flow passages (compressor disk with straight passages, with backward swept passages and with forward swept passages) were studied and effect of inlet flow angle on pressure loss characteristics in the intermediate shaft were examined. The experiment results indicated that, comparing to the straight passages, the backward swept passages realize a major decrease of pressure loss in the intermediate shaft. On the other hand, in the case of the forward swept passages, a rapid increase of pressure loss with decreasing the Rotation number was observed

Personalized Treatment for Infantile Ascending Hereditary Spastic Paralysis Based on In Silico Strategies

Infantile onset hereditary spastic paralysis (IAHSP) is a rare neurological disease diagnosed in less than 50 children worldwide. It is transmitted with a recessive pattern and originates from mutations of the ALS2 gene, encoding for the protein alsin and involved in differentiation and maintenance of the upper motoneuron. The exact pathogenic mechanisms of IAHSP and other neurodevelopmental diseases are still largely unknown. However, previous studies revealed that, in the cytosolic compartment, alsin is present as an active tetramer, first assembled from dimer pairs. The C-terminal VPS9 domain is a key interaction site for alsin dimerization. Here, we present an innovative drug discovery strategy, which identified a drug candidate to potentially treat a patient harboring two ALS2 mutations: one truncation at lysine 1457 (not considered) and the substitution of arginine 1611 with a tryptophan (R1611W) in the C-terminus VPS9. With a protein modeling approach, we obtained a R1611W mutant model and characterized the impact of the mutation on the stability and flexibility of VPS9. Furthermore, we showed how arginine 1611 is essential for alsin’s homo-dimerization and how, when mutated to tryptophan, it leads to an abnormal dimerization pattern, disrupting the formation of active tetramers. Finally, we performed a virtual screening, individuating an already therapy-approved compound (MK4) able to mask the mutant residue and re-establishing the alsin tetramers in HeLa cells. MK4 has now been approved for compassionate use