intraventricular-transplantation-engineered-neuronal-precursors-for

Intraventricular Transplantation of Engineered Neuronal Precursors for In Vivo Neuroarchitecture Studies

Gene control of neuronal cytoarchitecture is currently the subject of intensive investigation. Described here is a simple method developed to study in ...

SISSA

This chapter will focus on molecular mechanisms governing early steps of cortico-cerebral arealization. This is a very complex and intricate field, in which a great number of experimental investigations have been performed and vigorous scientific debates have taken place in recent years. We will try to provide an organic synthesis of this subject, summarizing the main experimental results and their theoretical implications.

Molecular bases of cortico-cerebral regionalization.

To promote expression of endogenous genes ad libitum, we developed a novel, programmable transcription factor prototype. Kept together via an MS2 coat protein/RNA interface, it includes a fixed, polypeptidic transactivating domain and a variable RNA domain that recognizes the desired gene. Thanks to this device, we specifically upregulated five genes, in cell lines and primary cultures of murine pallial precursors. Gene upregulation was small, however sufficient to robustly inhibit neuronal differentiation. The transactivator interacted with target gene chromatin via its RNA cofactor. Its activity was restricted to cells in which the target gene is normally transcribed. Our device might be useful for specific applications. However for this purpose, it will require an improvement of its transactivation power as well as a better characterization of its target specificity and mechanism of action. 

Upregulating endogenous genes by an RNA-programmable artificial transactivator.

Small activating RNAs (saRNAs), targeting endogenous genes and stimulating their transcription, are a promising tool for implementing a variety of neurotherapeutic strategies. Among these there is the stimulation of select histogenetic subroutines for purposes of cell-based brain repair, as well as the therapeutic treatment of gene expression deficits underlying severe neurological disorders.We employed RNA activation (RNAa) to transactivate the Emx2 transcription factor gene in embryonic cortico-cerebral precursor cells. This led to enhanced self-renewal, delayed differentiation, and reduced death of neuronally committed precursors, resulting in a remarkable expansion of the neuronogenic precursors pool. These results are of paramount interest for purposes of gene-promoted brain repair. As such, RNAa makes therapeutic stimulation of neuronogenesis via Emx2 overexpression a feasible goal, preventing the drawbacks of exogenous gene copies introduction.Moreover, we employed RNAa to achieve a gentle transactivation of the Foxg1 transcription factor gene, specifically in cortico-cerebral cells. This manipulation led to an appreciable biological outcome, while complying with endogenous gene tuning linked to early central nervous system regionalization and late activity of neocortical projection neurons. Foxg1-activating miRNAs stimulated RNApolII recruitment, possibly via Ago1. One of them worked promisingly in vivo. As such, RNAa can be a valuable approach for therapeutic treatment of the FOXG1-haploinsufficiency-linked variant of the Rett syndrome. Remarkably, hemizygosity for specific genes and polygenic chromosomal segments underlies a huge number of neuropathological entities for which no cure is presently available. Based on the results reported above, RNAa might be a simple and scalable approach for fixing this class of problems.

Article	Year
Molecular bases of cortico-cerebral regionalization. Progress in brain research\| PubMed ID: 21489382	2011
Upregulating endogenous genes by an RNA-programmable artificial transactivator. Nucleic acids research\| PubMed ID: 26152305	2015
Enhancing Neuronogenesis and Counteracting Neuropathogenic Gene Haploinsufficiencies by RNA Gene Activation. Advances in experimental medicine and biology\| PubMed ID: 28639189	2017

Antonello Mallamaci

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