Table 2.

The Summary of Current Examinations of Links Between miRNA Regulation and TREDs

Disease	miRNA change/regulation	miRNA target prediction algorithms	Methods used for experimental validation	Experimental models	References
Spinocerebellar ataxia type 1 (SCA1)	miR-19, -101 and -130a downregulate ATXN1 gene	PicTar was used to computationally predict miRNAs targeting ATXN1 transcript. Eight most likely miRNAs were chosen for experimental validation	transfection with miRNA duplexes and their specific inhibitors followed by western blot analysis and RT-PCR	MCF7, HEK293T, NIH3T3 and HeLa cell lines	[111]
			luciferase reporter assays with vectors carrying 3’UTR fragments or mutated target sites	HeLa cell line
			miRNA detection by northern blot analysis and in situ hybridization of mouse RNAs derived from cerebellum	C57/B6 WT mouse
			cell death assays with mutant ATXN1deprived of target sites	HEK293T cell line
Spinocerebellar ataxia type 3 (SCA3) and possibly other polyQ disorders	ban, a dma-miRNA, modulates polyQ-toxicity	–	phenotype comparison analysis (mutants)	D. melanogaster	[118, 119]
			cell death assays	D. melanogaster cell line with ban overexpression
			Dicer downregulation	flies and human cell lines
Dentatorubral pallidoluysian atrophy (DRPLA)	dma-miR-8 downregulates D. melanogaster atrophin gene	–	phenotype comparison analysis (mutants)	D. melanogaster	[129]
			mutant expression profiling microarray analysis	fruit fly pupae
			real-time RT-PCR with intron specific primers	fruit fly pupae
			luciferase reporter assay with vectors containing 3’UTR with mutated target sites	fruit fly pupae and S2 cell line
			in vivo studies of miR-8/atrophin functionality (death assays of mutants with atrophin and/or miR-8 underexpression, immmunocytochemistry for apoptotic cell detection)	D. melanogaster, fruit fly embryos
Huntington’s Disease (HD)	downregulation of miR-9, -9*, -29b, -124 and upregulation of miR-132 associates with HD	–	quantitative real-time PCR (qPCR) using TaqMan miRNA assays	human brain post mortem samples of the Brodmann’s area 4 (BA4) cortex	[130]
			co-transfection of miRNA precursors and REST/CoREST 3’UTRs with luciferase assay followed by western blot analysis	HEK293 cell line
Huntington’s Disease (HD)	downregulation of miR-132 and upregulation of miR-29a and -330 associate with HD	–	infection with adenovirus expressing a dominant-negative REST construct followed by RT PCR	cell lines of wt and mutant Hdh knock-in embryonic mice	[131, 132]
			qPCR using pre-miRNA stem loop primers	R6/2 mouse (and human) post mortem samples of the cortex (BA4 cortex)
Huntington’s Disease (HD)	downregulation of 15 miRNAs1 and upregulation of 192 and miRNA editing alterations associate with HD	microPred pipeline for novel miRNAs prediction [133], TargetScan for prediction of genes regulated by altered miRNAs	massively parallel sequencing of small non-coding RNAs (ncRNAs) followed by TaqMan microRNA assays	human brain post mortem samples of the frontal cortex (FC) and the striatum (ST)	[127]
Huntington’s Disease (HD)	downregulation of 15 miRNAs3 and upregulation of 94 associates with HD	miRNAMap 2.0 resource [77] (miRanda, TargetScan, RNAHybrid) was used for prediction of genes regulated by altered miRNAs	qPCR using stem loop primers	STHdhQ111/HdhQ111 cells - cell lines of wt and mutant Hdh knock in embryonic mice	[115]
Spinocerebellar ataxia type 17 (SCA17)	miR-146a downregulates TBP gene		luciferase reporter assay with vectors containing exogenous TBP 3’UTR with western blot analysis co-transfection of miRNA precursors and TBP 3’UTR followed by northern blot analysis
Myotonic dystrophy type 1 (DM1)	overexpression of miR-206 associates with DM1	–	qPCR using TaqMan microRNA assays northern and western blot analysis in situ hybridization of miR-206 using locked nucleic acid probes (LNA)	human muscle samples of vastus lateralis	[134]
Fragile X syndrome (FXS)	dFmrp associates with RISC and endogenous miRNAs	–	transcfection with vectors containing dFmrp co-immunoprecipitation with RISC proteins and miRNAs, norther and western blot analysis	S2 cell line	[123]
Fragile X syndrome (FXS)	dFmrp is required for processing of miR-124a	–	in situ hybridization of dma-miR-124a	D. melanogaster embryos	[125]
			immunoprecipitation with miR-124a, norther and western blot analysis qPCR using stem loop primers	transgenic fruit fly pupae
Fragile X syndrome (FXS)	miR-19b, -302b* and -323-3p downregulate FMR1 gene	intersection of computationally predicted targets by miRbase [3], miRanda and miRDB was used for validation	luciferase reporter assays with vectors containing native 3’UTR or with mutated target sites combined with co-transfection of GFP-tagged plasmids expressing pre-miRNAs	HEK293 cell line	[116]

¹miR-95, -124, -128, -127-3p, -139-3p, -181d, -221, -222, -382, -383, -409-5p, -432 , -433, -485-3p and -485-5p.

²miR-15b, -16, -17, -19b, -20a, -27b, -33b, -92a, -100, -106b, -148b, -151-5p, -193b, -219-2-3p, -219-5p, -363, -451, -486-5p and -887.

³miR-9, -9*, -100, -125b, -135a, -135b,-138, -146a, -150, -181c, -190, -218, -221, -222 and -338-3p.

⁴miR-145, -199-5p, -199-3p, -148a, -127-3p, -200a, -205, -214 and -335-5p.