Actin is an important protein in nucleus and has been implicated in transcription, however, the mechanism of its function in transcription is still not clear. In this article, we studied the role of actin in the regulation of human CSF1 gene transcription. Our results showed that nuclear actin stimulates the activity of CSF1 promoter, and the role in augmenting CSF1 gene transcription requires the formation of chromatin and Z-DNA structure. The ATP binding motifs of nuclear actin are essential for its function in regulating CSF1 gene transcription, and upon actin overexpression, there is an increase in the ATPase activity of nuclear proteins. Further investigation revealed that nuclear actin regulates CSF1 gene transcription in a BRG1 independent manner. Together, these original results have provided evidence for further understanding the mechanism of nuclear actin in regulating gene transcription.