Molecular cloning and characterization of cathepsin F gene from olive flounder Paralichthys Olivaceus

Abstract

We isolated a homologue of cathepsin F from cDNA library of olive flounder liver. A 2,077 kb full-length cDNA encoding a predicted polypeptide of 474 amino acids was sequenced. The flounder cathepsin F exhibits a domain structure typical for papain-like cysteine proteases, a 17 amino acid N-terminal hydrophobic signal sequence followed by an extraordinarily long propeptide of 244 amino acids and the domain of the mature protease comprising 213 amino acids. The mature region contains all features characteristic of a papain-like cysteine protease, including the highly conserved cysteine, histidine and asparagine residues of the 'catalytic triad'. The cathepsin F protein showed 49−99% amino acid sequence identity with other known cathepsin F sequences. An in vivo expression study showed that cathepsin F mRNA was expressed predominantly in brain, liver, eye and heart, and moderately in other tissues. The accumulation of cathepsin F mRNA in early stage of development increased with development. This expression pattern suggests that flounder cathepsin F has been implicated in the growth and reproduction regulation.

Expression Analysis of Cathepsin F during Embryogenesis and Early Developmental Stage in Olive Flounder (Paralichthys olivaceus)

“…In previous study, Kim et al ( 2010 ) reported cathepsin F that isolated from olive flounder liver cDNA library was evenly distributed all tissues in adult flounder and transcriptional level was gradually increased from 7 dph to 34 dph. But, Fabra & Cerdà ( 2004 ) described that cathepsin F mRNA was found throughout the oocyte growth or vitellogenesis period and a transient accumulation during oocyte maturation in teleost Fundulus heteroclitus .…”

“…The homologs search of the EST sequence with the corresponding amino acid sequence was examined using blast searches of Genbank and sequence alignment was performed with the CLUSTALW program ( ). A databank search showed that liver-1-D2 was revealed to be homologous with published flounder cathepsin F ( Kim et al, 2010 ) in olive flounder and has significant high identity above 98%. When compared to cathepsin F of other species were 98% with olive flounder (FJ436098.1), 91% with zebra mbuna (XP_004563245.1), 91% with nile tilapia (XP_003451852.1), 89% with marine medaka (AEA51106.1), 88% with atlantic salmon (ACN10907.1), 88% with ricefish (XP_004073613.1) and 87% with zebrafish (NP_001071036.1) (Fig.…”

“…To investigate the tissue specific expression of the cystein protease, oligonucleotide primers were designed according to previously isolated cathepsin F gene in olive flounder ( Kim et al, 2010 ) and then we examined by RT-PCR using isolated various tissues from 2-year-old olive flounder. Transcriptional level of cathepsin was compared between the tissues with beta-actin as the internal gene.…”

“…2 ). The recent study was demonstrated that cathepsin F was ubiquitously expressed in most of the tested tissues by RT-PCR ( Ahn et al, 2009 ; Kim et al, 2010 ). In this study, to analyze a more accurate patterns of tissue specific expression we examined the tissue distribution of cathepsin F using various tissues from healthy olive flounder of 90 days, 10 month, and 2 year aged.…”

“…In previous studies, olive flounder cathepsin F gene has been only reported that molecular cloning, enzymatic characterization and gene expression during immune responses ( Ahn et al, 2009 ; Kim et al, 2010 ), but the gene expression patterns for early developmental stage after fertilization was unknown. Therefore, the aim of this study was to investigate the gene expression patterns of cathepsin F in various tissues of adult as well as different developmental stages.…”