Aberrant gene expression is a hallmark of prostate cancer (PCa), the second deadliest disease affecting males worldwide. Dysregulation of miRNA has been associated with the progression of PCa and in recent studies, miRNA 574-3p was found to be upregulated in cancerous prostate tissue. In this study, we characterize the effects of upregulated miRNA 574-3p on gene expression in the tumor microenvironment through different bioinformatic tools such as Diana-Tools, the KEGG Pathway Database, and the Reactome Database. We have identified nine regulatory genes that are targeted by miRNA 574-3p and downregulated in prostate cells. Pathway analysis of these genes shows that they are involved in the regulation of the Notch signaling pathway, Wnt signaling pathway, apoptosis, DNA damage response, G1 to S cell-cycle control, inflammatory response pathway, angiogenesis, translation factors, and the expression of oncogenes. Our results show the oncogenic potential of miRNA 574-3p in PCa progression and metastasis. Moreover, this study highlights the complex molecular mechanisms and pathways affected by the upregulation of miRNA 574-3p in prostate cells. In future studies, the presented data may aid in designing new therapies for PCa with improved efficacy.