Depletion of epsilon-COP in the COPI Vesicular Coat Reduces Cleistothecium Production in Aspergillus nidulans

We have previously isolated epsilon-COP, the alpha-COP interactor in COPI of Aspergillus nidulans, by yeast two-hybrid screening. To understand the function of epsilon-COP, the aneA+ gene for epsilon-COP/AneA was deleted by homologous recombination using a gene-specific disruption cassette. Deletion of the epsilon-COP gene showed no detectable changes in vegetative growth or asexual development, but resulted in decrease in the production of the fruiting body, cleistothecium, under conditions favorable for sexual development. Unlike in the budding yeast Saccharomyces cerevisiae, in A. nidulans, over-expression of epsilon-COP did not rescue the thermo-sensitive growth defect of the alpha-COP mutant at 42degrees C. Together, these data show that epsilon-COP is not essential for viability, but it plays a role in fruiting body formation in A. nidulans.

Genomic Organization of ancop Gene for alpha-COP Homolog from Aspergillus nidulans

We have cloned a alpha-COP homolog, ancop, from Aspergillus nidulans by colony hybridization of chromosome specific library using alpha-COP homologous fragment as a probe. The probe DNA was amplified with degenerated primers designed by comparison of conserved region of the amino acid sequences of Saccharomyces cerevisiae alpha-COP, Homo sapiens HEP-COP, and Drosophila melanogaster alpha-COP. Full length cDNA clone was also amplified by RT-PCR. Comparison of genomic DNA sequence with cDNA sequence obtained by RT-PCR revealed 7 introns. Amino acid sequence similarity search of the anCop with other alpha-COPs gave an overall identity of 52% with S. cerevisiae, 47% with human and bovine, 45% with Drosophila and Arabidopsis . In upstream region from the transcription start site, a putative TATA and CAAT motif were also identified.