Post-Transcriptional Regulation of the Huntingtin Gene by MicroRNAs and PIWI-Interacting RNAs in Neurodevelopment and Neurodegenerative Processes

Huntington’s disease (HD) is a neurodegenerative disorder caused by cytosine–adenine–guanine (CAG) repeat expansion in the huntingtin (HTT) gene, which plays critical roles in both neuronal survival and early neurodevelopment. Small non-coding RNAs, including microRNAs (miRNAs) and PIWI-interacting RNAs (piRNAs), are key regulators of post-transcriptional gene expression. This study aimed to systematically investigate miRNA- and piRNA-mediated regulation of HTT mRNA and associated functional gene networks. HTT mRNA sequences were obtained from GenBank, while miRNA and piRNA sequences were retrieved from miRBase and piRBase. Binding sites (BSs) were predicted using the MirTarget program. A total of 195 miRNA BSs corresponding to 140 miRNAs were identified, predominantly within the coding sequence (CDS). Among these, 34 thermodynamically stable binding sites (ΔG ≤ −100 kJ/mol), involving 23 miRNAs, were identified. Network analysis revealed a neuronal module centered on mechanistic target of rapamycin (MTOR), B-cell lymphoma 2 (BCL2), synapsin I (SYN1), calcium/calmodulin-dependent protein kinase II alpha (CAMK2A), tubulin alpha 1a (TUBA1A), and huntingtin-associated protein 1 (HAP1). Analysis of 164 999 piRNAs identified 536 predicted binding sites corresponding to 207 unique piRNAs, most of which were localized within the coding region (93.8%). Binding energies reached −168 kJ/mol, with an average of −129 kJ/mol. Highly recurrent interactions were observed for piR-70378, piR-32860, piR-100200, and piR-144492. piRNA-associated targets included genes involved in developmental processes, including distal-less homeobox 6 (DLX6), E2F transcription factor 4 (E2F4), SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 2 (SMARCA2), DMRT-like family A2 (DMRTA2), and Zic family member 5 (ZIC5). Importantly, we propose a stage-specific regulatory model in which piRNAs predominantly contribute to early neurodevelopmental processes, whereas miRNAs mainly regulate neuronal function and survival in differentiated cells. These findings suggest that miRNAs and piRNAs regulate complementary neuronal and developmental pathways, highlighting a previously underexplored dual-layer regulatory mechanism of HTT and providing a conceptual framework for future biomarker discovery and therapeutic strategies in HD.