Bitondi Márcia M G, Nascimento Adriana M, Cunha Adriana D, Guidugli Karina R, Nunes Francis M F, Simões Zilá L P
Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Brazil.
Arch Insect Biochem Physiol. 2006 Oct;63(2):57-72. doi: 10.1002/arch.20142.
An N-terminal amino acid sequence of a previously reported honey bee hexamerin, HEX 110 [Danty et al., Insect Biochem Mol Biol 28:387-397 (1998)], was used as reference to identify the predicted genomic sequence in a public GenBank database. In silico analysis revealed an ORF of 3,033 nucleotides that encompasses eight exons. The conceptual translation product is a glutamine-rich polypeptide with a predicted molecular mass of 112.2 kDa and pI of 6.43, which contains the conserved M and C hemocyanin domains. Semiquantitative and quantitative RT-PCR with specific primers allowed for an analysis of mRNA levels during worker bee development and under different physiological conditions. Concomitantly, the abundance of the respective polypeptide in the hemolymph was examined by SDS-PAGE. Hex 110 transcripts were found in high levels during the larval stages, then decreased gradually during the pupal stage, and increased again in adults. HEX 110 subunits were highly abundant in larval hemolymph, decreased at the spinning-stage, and remained at low levels in pupae and adults. In 5th instar larvae, neither starvation nor supplementation of larval food with royal jelly changed the Hex 110 transcript levels or the amounts of HEX 110 subunit in hemolymph. In adult workers, high levels of Hex 110 mRNA, but not of the respective subunit, were related to ovary activation, and also to the consumption of a pollen-rich diet.
先前报道的蜜蜂六聚体蛋白HEX 110的N端氨基酸序列[丹蒂等人，《昆虫生物化学与分子生物学》28:387 - 397(1998)]被用作参考，以识别公共基因库数据库中的预测基因组序列。电子分析显示一个3033个核苷酸的开放阅读框，包含8个外显子。概念翻译产物是一种富含谷氨酰胺的多肽，预测分子量为112.2 kDa，pI为6.43，含有保守的M和C血蓝蛋白结构域。使用特异性引物进行的半定量和定量RT-PCR分析了工蜂发育过程中和不同生理条件下的mRNA水平。同时，通过SDS-PAGE检测血淋巴中相应多肽的丰度。发现Hex 110转录本在幼虫阶段含量很高，在蛹期逐渐下降，在成虫中再次升高。HEX 110亚基在幼虫血淋巴中含量很高，在吐丝阶段减少，在蛹和成虫中保持低水平。在5龄幼虫中，饥饿或用蜂王浆补充幼虫食物都不会改变Hex 110转录本水平或血淋巴中HEX 110亚基的含量。在成年工蜂中，高水平的Hex 110 mRNA，而非相应亚基，与卵巢激活以及富含花粉的饮食消耗有关。
