Analysis Pak Amphioxus COMMD4 Clone And Immune Function

COMMD4proteins are a group of recently discovered proteins which are characterized by the presence of a unique COMM domain (copper metabolism gene MURR1domain) of70to85amino acids in their extreme carboxy termini. The COMMD protein family has ten members (COMMD1-10) which share a carboxyl-terminal COMM domain. It has been showed that COMM domain could be able to bind to other factors such as Cul2, and plays a key role in inhibiting NF-κB. Little information is now available regarding the biochemical characteristics and functions of the other members, excepting COMMD1. Cephalochordate amphioxus is a kind of transitional species between invertebrate and vertebrate. It is well-known model organism widely used for interspecies comparative genome studies and developmental homology analysis. With the Branchiostoma floridae genome sequencing, the attention has been focused on amphioxus immune increasingly.In this study, we cloned the full length of AmphiCOMMD4gene. Full length of AmphiCOMMD4cDNA of1008bp was identified and an open reading frame (ORF) with603bp was predicted. Besides the ORF, AmphiCOMMD4cDNA contained a5’-UTR with llbp and394bp3’-UTR including a poly (A) tail with12bp and a canonical polyadenylation signal (AATAAA). The ORF encoded a putative AmphiCOMMD422kDa protein containing200amino acids. The AmphiCOMMD4protein shared55.5-82.5%similarity and38.8-64.0%identity to other known COMMD4homolog. The phylogenetic tree constructed using the sequeneces of representative COMMD proteins including AmphiCOMMD4demonstrated that AmphiCOMMD4was clustered with COMMD4proteins, separating from all the other9family members. The phylogenetic tree constructed using the sequeneces of representative COMMD4proteins including AmphiCOMMD4suggested that the AmphiCOMMD4was positioned at the bottom of the chordate, which was in general accordance with the classical taxonomy. The analysis of the genomic sequence of AmphiCOMMD4showed that only one exon was identified in the genomic sequence of AmphiCOMMD4. This pattern is consistent with that from sea squirt, but different from those from fruitfly, sea urchin and vertebrates. Intron gain or retention in sea urchin and vertebrate to a certain extent may be related to the function of introns. However, it is unknown whether six intron-loss events were generated in COMMD4gene of amphioxus and sea squirt by retroposition and homologous recombination. Conservation analysis of the ten COMMD family members in eight species suggested that different species contained different numbers of COMMD proteins. Vertebrates, for example, had all the ten COMMD family members. The amphioxus may have secondarily lost COMMD1, and the absence of these genes may be also due to the incomplete genome assembly. It is found that the AmphiCOMMD4was ubiquitously and differentially expressed in five investigate tissue (muscles, gills, intestine, hepatic cecum and gonad), but varied expression levels were observed among the different tissues with the highest level in gonad tissue. The expression level of AmphiCOMMD4gene was induced by LPS stimulation. It first increased, then decreased (expression levels of highest and lowest values are found at the6h and12h, respectively), and returned to normal level at48h after LPS stimulation, In conclusion, our study elaborates the COMMD4gene in amphioxus and provided an insight into the innate immunity and the evolution of the COMMD protein family.