Proteomic identification of intracellular vesicle trafficking and protein glycosylation requirements for lumen inflation in Ciona notochord.

Lumen formation and inflation are crucial steps for tubular organ morphogenesis, yet the underling mechanism remains largely unrevealed. Here, we applied 4D proteomics to screen the lumenogenesis-related proteins and revealed the biological pathways potentially that are involved in lumen inflation during notochord lumen formation in the ascidian Ciona savignyi. In total, 910 differentiated expressed proteins (DEPs) were identified before and after notochord lumen formation utilizing Mfuzz analysis. Those DEPs were grouped into four upregulated clusters based on their quantitative expression patterns; the functions of these proteins were enriched in protein metabolic and biosynthetic process, the establishment of localization, and vesicle-mediated transport. We analyzed the vesicle trafficking cluster and focused on several vesicle transport hub proteins. In vivo function-deficient experiments showed that mutation of vesicle transport proteins resulted in an abnormal lumen in notochord development, demonstrating the crucial role of intracellular trafficking for lumen formation. Moreover, abundant extracellular matrix proteins were identified, the majority of which were predicted to be glycosylated proteins. Inhibition of glycosylation markedly reduced the lumen expansion rate in notochord cells, suggesting that protein glycosylation is essential for lumenogenesis. Overall, our study provides an invaluable resource and reveals the crucial mechanisms in lumen formation and expansion.

Temporospatial hierarchy and allele-specific expression of zygotic genome activation revealed by distant interspecific urochordate hybrids.

Zygotic genome activation (ZGA) is a universal process in early embryogenesis of metazoan, when the quiescent zygotic nucleus initiates global transcription. However, the mechanisms related to massive genome activation and allele-specific expression (ASE) remain not well understood. Here, we develop hybrids from two deeply diverged (120 Mya) ascidian species to symmetrically document the dynamics of ZGA. We identify two coordinated ZGA waves represent early developmental and housekeeping gene reactivation, respectively. Single-cell RNA sequencing reveals that the major expression wave exhibits spatial heterogeneity and significantly correlates with cell fate. Moreover, allele-specific expression occurs in a species- rather than parent-related manner, demonstrating the divergence of cis-regulatory elements between the two species. These findings provide insights into ZGA in chordates.

Combined treatment of neurotrophin-3 gene and neural stem cells is ameliorative to behavior recovery of Parkinson's disease rat model.

Neural stem cell transplantation therapy was developed for replacing lost or damaged neural cells for the neurodegenerative disease, including Parkinson's disease (PD), in which dopaminergic neuron cells are lost. The growth factor, neurotrophin-3(NT-3), has been shown to promote neuroregeneration, differentiation and migration during brain development. In this report, we construct rat neural stem cells that express neurotrophin-3 endogenously (rNSC-NT3) and transplant them into 6-hydroxydopamine (6-OHDA)-treated Parkinsonian rats. Molecular approaches including quantitative real time PCR, Western blot and immunocytochemistry were used to identify the expression of NT-3 and the differentiation of planted cells. Behavioral recover was also tested. The result indicated that combined treatment of neurotrophin-3 gene and neural stem cells had a functional impact on reversing the main symptoms of the Parkinson's disease that significantly reduced apomorphine-induced rotational asymmetry and improved spatial learning ability. The rNSCs-NT3 is able to differentiate into dompaminergic neuron in the ventral tegmental area (VTA) and the medial forebrain bundle (MFB), and migrated around the lesion site. Endogenous expressed NT-3 exerts induction and trophic effects on neural stem cells. The rNSCs-NT3 showed higher activity than the rNSCs in regenerating tyrosine hydroxylase positive cell numbers and migrating distance, behavior improving in this dopa-deficit rat model. These findings suggest that the neural stem cells expressed NT-3 endogenously would be a better graft candidate for the treatment of Parkinson's disease.