Abstract
Non-coding RNA, including microRNA (miRNA) serves critical regulatory functions in the developing brain. The let-7 family of miRNAs has been shown to regulate neuronal differentiation, neural subtype specification, and synapse formation in animal models. However, the regulatory role of human let-7c (hsa-let-7c) in human neuronal development has yet to be examined. Let-7c is encoded on chromosome 21 in humans and therefore may be overexpressed in human brains in Trisomy 21 (T21), a complex neurodevelopmental disorder. Here, we employ recent developments in stem cell biology to show that hsa-let-7c mediates important regulatory epigenetic functions that control the development and functional activity of human induced neuronal cells (iNs). We show that overexpression of hsa-let-7c in human iNs derived from induced pluripotent stem (iPS), as well as embryonic stem (ES), cells leads to morphological as well as functional deficits including impaired neuronal morphologic development, synapse formation and synaptic strength, as well as a marked reduction of neuronal excitability. Importantly, we have assessed these findings over three independent genetic backgrounds, showing that some of these effects are subject to influence by background genetic variability with the most robust and reproducible effect being a striking reduction in spontaneous neural firing. Collectively, these results suggest an important function for let-7 family miRNAs in regulation of human neuronal development and raise implications for understanding the complex molecular etiology of neurodevelopmental disorders, such as T21, where let-7c gene dosage is increased.
Highlights
Synaptic transmission mediates fast information flow in the brain
We investigated the role of hsa-let-7c in human neuronal and synaptic development by overexpressing the mature let-7c sequence in pluripotent stem cell-derived induced neuronal cells
We have discovered that let-7c overexpression leads to a number of deficits in neuronal morphology, electrical excitability, and synaptic transmission, the most prominent being a striking reduction in spontaneous neuronal firing
Summary
Synaptic transmission mediates fast information flow in the brain. As such, synapses represent the fundamental unitary elements of neuronal circuits and brain network systems that govern all behavior and cognitive processes. We found that lentiviral-mediated over expression of hsa-let-7c impairs neuronal and synaptic development and markedly reduces neuronal excitability We assessed these functions over three different pluripotent cell lines derived from independent genetic backgrounds, revealing a consistent role for let-7c in regulating spontaneous neuronal firing activity. These analyses show that some features of epigenetic regulation by let-7c may be subject to regulation by genetic background or epigenetic state, as only two of the lines analyzed showed a statistically significant synaptic phenotype and intrinsic excitability deficit These results implicate let-7c as an important regulator in human neuronal development and function, and shed light onto possible epigenetic regulatory mechanisms associated with gene-dosage dependent pathophysiology mediated by non-coding RNAs associated with T21 in humans
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