Impaired myoblast differentiation and muscle IGF-1 receptor signaling pathway activation after N-glycosylation inhibition.

The role of N-glycosylation in the myogenic process remains poorly understood. Here, we evaluated the impact of N-glycosylation inhibition by Tunicamycin (TUN) or by phosphomannomutase 2 (PMM2) gene knockdown, which encodes an enzyme essential for catalyzing an early step of the N-glycosylation pathway, on C2C12 myoblast differentiation. The effect of chronic treatment with TUN on tibialis anterior (TA) and extensor digitorum longus (EDL) muscles of WT and MLC/mIgf-1 transgenic mice, which overexpress muscle Igf-1Ea mRNA isoform, was also investigated. TUN-treated and PMM2 knockdown C2C12 cells showed reduced ConA, PHA-L, and AAL lectin binding and increased ER-stress-related gene expression (Chop and Hspa5 mRNAs and s/uXbp1 ratio) compared to controls. Myogenic markers (MyoD, myogenin, and Mrf4 mRNAs and MF20 protein) and myotube formation were reduced in both TUN-treated and PMM2 knockdown C2C12 cells. Body and TA weight of WT and MLC/mIgf-1 mice were not modified by TUN treatment, while lectin binding slightly decreased in the TA muscle of WT (ConA and AAL) and MLC/mIgf-1 (ConA) mice. The ER-stress-related gene expression did not change in the TA muscle of WT and MLC/mIgf-1 mice after TUN treatment. TUN treatment decreased myogenin mRNA and increased atrogen-1 mRNA, particularly in the TA muscle of WT mice. Finally, the IGF-1 production and IGF1R signaling pathways activation were reduced due to N-glycosylation inhibition in TA and EDL muscles. Decreased IGF1R expression was found in TUN-treated C2C12 myoblasts which was associated with lower IGF-1-induced IGF1R, AKT, and ERK1/2 phosphorylation compared to CTR cells. Chronic TUN-challenge models can help to elucidate the molecular mechanisms through which diseases associated with aberrant N-glycosylation, such as Congenital Disorders of Glycosylation (CDG), affect muscle and other tissue functions.

Role of melanin in the black yeast fungi Aureobasidium pullulans and Zalaria obscura in promoting tolerance to environmental stresses and to antimicrobial compounds.

The role of melanin in Aureobasidium pullulans ATCC 15233 and Zalaria obscura LS31012019, under simulated osmotic, oxidative, and high temperature stress conditions, on the susceptibility to essential oils (EOs) or antifungals and on the resistance to UV-C radiation was investigated. 93.6% of melanized A. pullulans and 92% of Z. obscura survived to 40 °C for 1 h compared to 77% and 76% of the non-melanized ones, while both yeasts tolerated a high concentration of NaCl (up to 30%) and H2O2 (up to 400 mM) regardless of melanin production. Higher EOs antifungal efficacy was observed in non-melanized cells (growth inhibition zone >30 mm) compared to the melanized ones (25 mm). Similarly, the lowest Minimum Inhibitory Concentrations (MIC) and Minimum Fungicidal Concentration (MFC) values were evidenced for Fluconazole, Clotrimazole, Bifonazole and Amphotericin in the non-melanized fungi. Increasing UV-C intensity (up to 2004.5 J/m2) caused total death in the non-melanized strains compared to about 30% growth reduction in the melanized ones. The results of this investigation, the first focused on the biological role of melanin in "black-fungi", are novel and encourage a better understanding of the biochemical features of melanin in the environmental adaptive ability of the new species Z. obscura.