Actin-related proteins (ARPs) share moderate sequence homology and basal structure with conventional actins and are found in all eukaryotes. While the functions of most of the divergent ARPs are not clear, several of them are localized to the nucleus and have been identified as components of various chromatin-modifying complexes. Using an antibody to Arabidopsis AtARP4, we found this conserved homolog of human BAF53 and yeast Arp4 is concentrated in the nucleoplasm of Arabidopsis, Brassica, and tobacco cells. To gain further insight into the role of ARP4, we have examined Arabidopsis plants that are defective in AtARP4 expression. Phenotypic analysis of the arp4-1 mutant allele, which has a T-DNA insertion in the promoter region and a moderate reduction in the level of AtARP4 protein expression, revealed partial sterility due to defects in anther development. Targeting the distinct, 3' UTR of AtARP4 transcripts with RNA interference caused a drastic reduction in the level of AtARP4 protein expression in several independent transgenic lines, and resulted in strong pleiotropic phenotypes such as altered organization of plant organs, early flowering, delayed flower senescence and high levels of sterility. Western blot analysis and immunolabeling demonstrated a clear correlation between reductions in the level of AtARP4 expression and severity of the phenotypes. Based on our results and data on the orthologs of AtARP4 in yeast and other organisms, we suggest that AtARP4 is likely to exert its effects on plant development through the modulation of chromatin structure and subsequent changes in gene regulation.