Abstract
UDP-N-acetylglucosamine pyrophosphorylases (UAP) function in the formation of extracellular matrix by producing N-acetylglucosamine (GlcNAc) residues needed for chitin biosynthesis and protein glycosylation. Herein, we report two UAP cDNA’s derived from two different genes (LmUAP1 and LmUAP2) in the migratory locust Locusta migratoria. Both the cDNA and their deduced amino acid sequences showed about 70% identities between the two genes. Phylogenetic analysis suggests that LmUAP1 and LmUAP2 derive from a relatively recent gene duplication event. Both LmUAP1 and LmUAP2 were widely expressed in all the major tissues besides chitin-containing tissues. However, the two genes exhibited different developmental expression patterns. High expression of LmUAP1 was detected during early embryogenesis, then decreased greatly, and slowly increased before egg hatch. During nymphal development, the highest expression of LmUAP1 appeared just after molting but declined in each inter-molting period and then increased before molting to the next stage, whereas LmUAP2 was more consistently expressed throughout all these stages. When the early second- and fifth-instar nymphs (1-day-old) were injected with LmUAP1 double-stranded RNA (dsRNA), 100% mortality was observed 2 days after the injection. When the middle second- and fifth-instar nymphs (3- to 4-day-old) were injected with LmUAP1 dsRNA, 100% mortality was observed during their next molting process. In contrast, when the insects at the same stages were injected with LmUAP2 dsRNA, these insects were able to develop normally and molt to the next stage successfully. It is presumed that the lethality caused by RNAi of LmUAP1 is due to reduced chitin biosynthesis of the integument and midgut, whereas LmUAP2 is not essential for locust development at least in nymph stage. This study is expected to help better understand different functions of UAP1 and UAP2 in the locust and other insect species.
Highlights
N-acetylglucosamine (GlcNAc), a substrate required for glycosylphosphatidylinositol (GPI) anchor formation, is an important component of glycosyl groups that modify extracellular proteins [1]
The full-length cDNA sequence of LmUAP1 is 2223 bp with an open reading frame (ORF) of 1455 bp and a 66-nucleotide 59untranslated region (UTR) and a 702-nucleotide 39-UTR, which encodes a protein of 484 amino acid residues
The full-length cDNA of LmUAP2 is 1995 bp with the ORF of 1482 bp and a 158nucleotide 59-UTR and a 355-nucleotide 39-UTR, which encodes a protein of 493 amino acid residues
Summary
N-acetylglucosamine (GlcNAc), a substrate required for glycosylphosphatidylinositol (GPI) anchor formation, is an important component of glycosyl groups that modify extracellular proteins [1]. GlcNAc contributes to the structure and function of various extracellular matrix. Chitin is an linear biopolymer of the sugar N-acetylglucosamine (GlcNAc) connected by b-1,4-linkages. Chitin is an integral part of cuticle, trachea and peritrophic matrix (PM) [2,3]. Insects must consistently synthesize and degrade chitin to allow ecdysis and regeneration of the PM. Current knowledge on insect chitin biosynthesis is still fragmentary
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