Canovas, M. | Search bioRxiv

bioRxiv2024

Maternal Immune Activation imprints translational dysregulation and differential MAP2 phosphorylation in descendant neural stem cells

Alterations induced by maternal immune activation (MIA) during gestation impact the subsequent neurodevelopment of progeny, a process that in humans, has been linked to the development of several neuropsychiatric conditions. To undertake a comprehensive examination of the molecular mechanisms governing MIA, we have devised an in vitro model based on neural stem cells (NSCs) sourced from fetuses carried by animals subjected to Poly I:C treatment. These neural progenitors demonstrate proliferative capacity and can be effectively differentiated into both neurons and glial cells. Transcriptomic, proteomic, and phosphoproteomic analyses conducted on these cellular models, in conjunction with counterparts from control treatments, revealed discernible shifts in the expression levels of a specific subset of proteins implicated in neuronal function. Noteworthy, we found an absence of congruence between these alterations at the transcriptomic level, suggesting that differences in protein translation contribute to the observed dysregulation. Furthermore, the phosphoproteomic data highlighted a discernible discrepancy in the basal phosphorylation of proteins between differentiated cells from both experimental groups, particularly within proteins associated with cytoskeletal architecture and synaptic functionality, notably those belonging to the MAP family. Observed alterations in MAP phosphorylation were found to potentially have functional consequences as they correlate with changes in neuronal plasticity and the establishment of neuronal synapses. Our data agrees with previous published observations and further underscore the importance of MAP2 phosphorylation state on its function and the impact that this protein has in neuronal structure and function.

neuroscience↗

bioRxiv2021

MODEL BASED OPTIMIZATION OF THE TERPENOIDS BIOSYNTHESIS IN E. coli

Terpenoids are a family of compounds with high industrial interest and the development of biotechnological production methods is essential to achieve more sustainable alternatives to traditional extraction and synthesis methods. The modification and engineering of the catalytic activity (kcat) have been shown to be a feasible strategy in the biotechnological realm. Accordingly, we introduce a novel optimization strategy based in the modification of the kcat of the enzymes and applied it to the maximization of the terpenoids synthesis in E. coli. This approach is fairly general and can be applied alone or in conjunction with classic optimization strategies such as the modification of enzymatic specific activities. For this purpose we first build up a reliable dynamic mathematical model of the alternative mevalonate pathway synthesis leading terpenoids biosynthesis in E. coli through the methyl-D-erythritol 4-phosphate (MEP) pathway. This model includes the 2-C-methyl-D-erythritol 2, 4-diphosphate (MEC) pumps that mediate MEC extracellular extrusion. Although the physiological significance of the MEC extrusion is still discussed and their biological function is not clear, we find that this process is a must to guarantee bacteria homeostasis and cell viability. We have identified the enzyme IspA as a bottleneck of the terpenoids biosynthesis, which is dual substrate for the two bisubstrate final reactions. Here are presented different ways to overcome this enzymatic flux restriction by modification of the IspA kcat or by introduction of new enzymes with parallel function. Our results show that the MEP pathway optimized solutions for kcat can yield a maximum of 17.68 fold increment in the terpenoids biosynthetic flux when the kcat are modified. This maximal solution involves the modification of 8 kcat and the corresponding Kms and Kds. Remarkably, this increase doesnt imply a change in the total enzyme concentration of the cell. This is favorable output since enzyme overproduction can compromise cell functionality. We also apply the overexpression of the enzyme activity approach and then we have compared and combined both strategies including scenarios as the deletion or import of the genes expressing enzymes not naturally present in E. coli. A combined strategy of enzymes concentrations and kcat modifications with changes up ti six enzyme levels allows a flux increment of 21.22 fold the basal value. It involves the incorporation of two IspA with similar GPP and IPP bisubstrate functions than the native one but low affinity for the DMAPP as substrate.

systems biology↗