microRNAs (miRNAs) are abundant, ∼21-nucleotide, noncoding regulatory RNAs. Each miRNA may regulate hundreds of mRNA targets, but the identities of these targets and the processes they regulate are poorly understood. Here we have explored the use of microarray profiling and functional screening to identify targets and biological processes triggered by the transfection of human cells with miRNAs. We demonstrate that a family of miRNAs sharing sequence identity with miRNA-16 (miR-16) negatively regulates cellular growth and cell cycle progression. miR-16-down-regulated transcripts were enriched with genes whose silencing by small interfering RNAs causes an accumulation of cells in G₀/G₁. Simultaneous silencing of these genes was more effective at blocking cell cycle progression than disruption of the individual genes. Thus, miR-16 coordinately regulates targets that may act in concert to control cell cycle progression.