Amphioxus allantoicase: molecular cloning, expression and enzymatic activity

Area of Science:

• Biochemistry
• Evolutionary Biology
• Chordate Genomics

Background:

• Allantoicase, an enzyme in purine metabolism, has undergone progressive truncation throughout chordate evolution.
• The precise evolutionary point at which allantoicase activity became phylogenetically extinct remains undetermined.

Purpose of the Study:

• To investigate the presence and activity of allantoicase in amphioxus (Branchiostoma belcheri tsingtauense), a primitive chordate.
• To determine the phylogenetic persistence of allantoicase function in early chordates.

Main Methods:

• Isolation of a cDNA encoding allantoicase from an amphioxus gut cDNA library.
• Reverse transcription polymerase chain reaction (RT-PCR) to analyze gene expression patterns in various amphioxus tissues.
• Enzyme activity assays to measure allantoicase function across different tissues.

Main Results:

• A 2441 bp cDNA encoding a 392-amino acid allantoicase protein was successfully isolated from amphioxus.
• RT-PCR revealed strong expression of amphioxus allantoicase in the hepatic caecum and weaker expression in other tissues.
• Enzyme activity assays confirmed high allantoicase activity in the hepatic caecum, with low or undetectable levels elsewhere.

Conclusions:

• Allantoicase is present and enzymatically active in the primitive chordate amphioxus.
• The hepatic caecum is the primary site of allantoicase expression and activity in amphioxus.
• These findings suggest that allantoicase function has persisted into early chordate lineages, challenging previous assumptions about its evolutionary extinction.