Aspergillus aculeatus secretes superior cellulolytic and hemicellulolytic enzymes on saccharification. It has been revealed that a Zn(II)2Cys6 transcription factor, XlnR, positively regulated those gene expressions in Aspergillus. However, we found that the xlnR gene disruption did not affect the transcription levels of cellobiohydrolase I (cbhI) and carboxymethylcellulase 2 (cmc2) genes in A. aculeatus. These data suggests that those genes are regulated by namely XlnR-independent signaling pathway. To identify regulator(s) participating in the regulation network, we established a positive screening system using a PcbhI-pyrG reporter fusion. Mutations were introduced by T-DNA insertion utilizing Agrobacterium tumefaciens-mediated transformation system, and which enabled to rescue the franks of T-DNA by PCR-based technique. Transformants were selected by indexing 5-FOA resistance and cellulose utilization deficiency, and then further analyzed on expression level of the cbhI and cmc2 by RT-PCR. One out of 6,000 transformants was, so far, identified as a cellulase induction-deficient mutant. The T-DNA was verified to insert into near 5"¢f-end of ORF encoding putative a Zn(II)2Cys6 transcription factor, namely factor A. The factor A gene disruption by homologous recombination resulted in reduction of the cbhI and cmc2 transcripts, demonstrating that Factor A participated in the XlnR-independent signaling pathway.