Department of Plant Pathology, Citrus Research and Education Center, Institute of Food and Agricultural Sciences, University of Florida, 700 Experiment Station Road, Lake Alfred, FL 33850, USA.
Curr Genet. 2010 Feb;56(1):43-51. doi: 10.1007/s00294-009-0278-2. Epub 2009 Dec 1.
Heterotrimeric G proteins play a profound role in the recognition and transduction of extracellular signals in eukaryotic cells. We characterized the AaG alpha1 gene, encoding a fungal Class I G alpha subunit of the GTP-binding protein, in Alternaria alternata of citrus. Interruption of AaG alpha1 with a marker gene resulted in fungal transformants producing fewer conidia, becoming hypersensitive to KCl, and displaying elevated lipolytic and pectolytic activities. Expression of a functional copy of AaG alpha1 in a null mutant restored all altered phenotypes to the wild type. The AaG alpha1 mutants, whose conidia germinate normally, caused necrotic lesions on citrus indistinguishable from wild type. Application of cAMP, its inhibitors (atropine and theophylline), or 3-isobutyl-1-methylxanthine (IBMX) decreased conidiation in the fungal strains carrying a functional AaG alpha1. In contrast, conidial formation in the null mutants was restored by dibutyryl-cAMP or by a low concentration of cAMP or theophylline (1 mM). Unlike the oxidative stress-responsive AaAP1 transcription activator or the AaFUS3 mitogen-activated protein kinase (MAPK), AaG alpha1 is not required for cellular resistance to oxidative, osmotic, or chemical stress. AaFUS3 has also been demonstrated to be essential for conidial formation, suggesting a possible interaction between AaG alpha1 and AaFUS3 during conidiation even though expression of AaG alpha1 was not affected by AaFUS3 and vice versa. Inactivation of AaG alpha1 suppressed accumulation of transcripts of the AaAP1 gene and the AaHSK1 gene encoding a histidine kinase. These are novel features that have not been previously characterized to be associated with the GTP-binding protein.
异三聚体 G 蛋白在真核细胞中外源信号的识别和转导中发挥着深远的作用。我们对柑橘青霉（Alternaria alternata）的 AaG alpha1 基因进行了特征描述，该基因编码真菌类 I G 蛋白的 GTP 结合蛋白的 alpha 亚基。用标记基因中断 AaG alpha1 的功能，导致真菌转化体产生的分生孢子减少，对 KCl 更加敏感，并显示出更高的脂解和果胶酶活性。在一个缺失突变体中表达一个有功能的 AaG alpha1 拷贝，可将所有改变的表型恢复为野生型。AaG alpha1 突变体的分生孢子正常萌发，但在柑橘上引起的坏死性病变与野生型无异。应用 cAMP、其抑制剂（阿托品和茶碱）或 3-异丁基-1-甲基黄嘌呤（IBMX）可减少携带功能性 AaG alpha1 的真菌菌株的分生孢子形成。相反，在缺失突变体中，二丁酰基-cAMP 或低浓度的 cAMP 或茶碱（1mM）可恢复分生孢子的形成。与氧化应激反应的 AaAP1 转录激活子或 AaFUS3 有丝分裂原激活蛋白激酶（MAPK）不同，AaG alpha1 不是细胞对氧化、渗透或化学应激的抗性所必需的。AaFUS3 也被证明是分生孢子形成所必需的，这表明在分生孢子形成过程中，AaG alpha1 和 AaFUS3 之间可能存在相互作用，尽管 AaG alpha1 的表达不受 AaFUS3 的影响，反之亦然。AaG alpha1 的失活抑制了 AaAP1 基因和编码组氨酸激酶的 AaHSK1 基因的转录本的积累。这些是以前尚未被认为与 GTP 结合蛋白有关的新特征。
