Small RNA and its application in andrology and urology
ABSTRACT Small non-coding RNAs such as small interfering RNA (siRNA), microRNA (miRNA) and piwi-interacting RNA (piRNA) exist in almost all kingdoms of organisms and have recently emerged as master regulators of gene expression to affect a diverse range of important biological processes. They exert their functions largely through two related but opposing mechanisms: RNA interference (RNAi) mediated by siRNA, miRNA and piRNA, and RNA activation (RNAa) mediated by small activating RNA (saRNA) and miRNA, leading to silencing and overexpression of target genes respectively. Dysregulation of these mechanisms have been implicated in a variety of human diseases including urological and andrological diseases. Importantly, both mechanisms can be readily harnessed for therapeutic purposes for a variety of diseases by using small RNA molecules as the "ribodrug". In this review, we highlight recent advances in the applications of small RNA as therapeutics for urological cancer, male infertile and erectile dysfunction.
- SourceAvailable from: Mahmoud Kandeel[Show abstract] [Hide abstract]
ABSTRACT: RNA interference (RNAi) is a highly specialized process of protein-siRNA interaction that results in the regulation of gene expression and cleavage of target mRNA. The PAZ domain of the Argonaute proteins binds to the 3' end of siRNA, and during RNAi the attaching end of the siRNA switches between binding and release from its binding pocket. This biphasic interaction of the 3' end of siRNA with the PAZ domain is essential for RNAi activity; however, it remains unclear whether stronger or weaker binding with PAZ domain will facilitate or hinder the overall RNAi process. Here we report the correlation between the binding of modified siRNA 3' overhang analogues and their in vivo RNAi efficacy. We found that higher RNAi efficacy was associated with the parameters of lower Ki value, lower total intermolecular energy, lower free energy, higher hydrogen bonding, smaller total surface of interaction and fewer van der Waals interactions. Electrostatic interaction was a minor contributor to compounds recognition, underscoring the presence of phosphate groups in the modified analogues. Thus, compounds with lower binding affinity are associated with better gene silencing. Lower binding strength along with the smaller interaction surface, higher hydrogen bonding and fewer van der Waals interactions were among the markers for favorable RNAi activity. Within the measured parameters, the interaction surface, van der Waals interactions and inhibition constant showed a statistically significant correlation with measured RNAi efficacy. The considerations provided in this report will be helpful in the design of new compounds with better gene silencing ability.PLoS ONE 02/2013; 8(2):e57140. DOI:10.1371/journal.pone.0057140 · 3.23 Impact Factor
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ABSTRACT: T-type calcium channels are a class of low voltage-dependent calcium channels that may be activated following minor depolarizations of the cell membrane. Cav3.1 is the dominant subtype of the T-type calcium channel in SH-SY5Y cells. T-type channels play a key role in the regulation of the intracellular calcium concentration, which is involved in the neurotoxic effect of local anesthetics. However, there is a lack of specific inhibitors of T-type calcium channels. The existing T-type calcium channel inhibitors exhibit poor specificity and may block the high voltage-dependent calcium channels, such as the L- and N-type channels. Furthermore, there is no selectivity to the subtype of the T-type calcium channel. Therefore, the development of a specific T-type calcium channel inhibitor may contribute to the elucidation of the functions and characteristics of T-type calcium channels. The aim of this study was to silence the Cav3.1 mRNA expression in SH-SY5Y cells via the RNA interference (RNAi) method in order to construct pshRNA-CACNA1G-SH-SY5Y cells and assess Cav3.1 mRNA and protein expression by western blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) to identify the constructed cell line. The results demonstrated that Cav3.1 mRNA and protein expression were significantly reduced following transfection with the SH-SY5Y cells by the supernatant liquors. The results also demonstrated that the pshRNA-CACNA1G-SH-SY5Y cells were successfully constructed. These findings may contribute to the elucidation of the functions of Cav3.1 in SH-SY5Y cells.07/2013; 1(4):669-673. DOI:10.3892/br.2013.117