Molecular cloning, characterization and gene expression analysis of aminolevulinic acid synthase in Litopenaeus...

5-Aminolevulinic acid synthase (ALAS) is the rate-limiting enzyme in the biosynthesis of heme, a prosthetic group that is found in hemoproteins, including those involved in molting. To better understand the roles of ALAS in L. vannamei (LvALAS), we analyzed its sequence and tissue distribution, the effects of age and bacterial infection on its gene expression, and the effects of LvALAS gene silencing. We also examined the expressions of three hemoproteins, the cytochrome oxidase subunit I (COX I) and subunit IV (COX IV) and catalase. Three LvALAS splicing variants were found in the hepatopancreas, with the main splicing variant having an open reading frame that encodes 532 aa. LvALAS transcripts were found in each of the eleven tissues tested in this study, with the highest gene expression in the intestine. The transcript abundances of LvALAS, COX I and COX IV in the hepatopancreas and stomach tended to decrease with age. LvALAS and catalase gene expressions significantly increased in the stomach after V. parahaemolyticus infection. LvALAS gene expression in the hepatopancreas, stomach and intestine (12- and 24-hours post-injection) was relatively lower in dsALAS-injected shrimp than in PBS-injected shrimp. All the PBS-injected shrimp molted after 8-10 days while no molting activity was observed in the dsALAS-injected shrimp group within the 14 days post-injection period. Our results provide evidence that (1) only the housekeeping form of ALAS exists in L. vannamei; LvALAS gene expression (2) decreases with age and (3) increases after bacterial infection; and (4) an ALAS-dependent pathway is necessary for proper molting in L. vannamei.