Lessons from a gene knockout mouse model: no phenotypes induced by deletion of the TPR4 domain of Ttc22

The TTC22 gene encodes a protein containing seven tetratricopeptide repeats (TPRs), which mediate protein‒protein interactions as chaperones. We previously reported that the level of TTC22 transcript variant 1 ( TTC22v1 ) was downregulated in human colon adenocarcinoma (COAD) and that TTC22 upregulated m6A-mediated WTAP and SNAI1 expression via the TTC22‒RPL4 interaction and subsequently promoted COAD metastasis. Thus, a commercially available C57BL/6N mouse model in which the Ttc22 exon 2&3 encoding the TPR4 (equal to the human TTC22 TPR3) domain was knocked out via CRISPR-Cas9 was used to evaluate the contribution of Ttc22 to the development of mice and COAD. Unfortunately, the long-term observation results demonstrated that Ttc22 knockout (KO, including Ttc22 ^-/+ or Ttc22 ^-/- ) did not affect the body weight, development, fertility, or spontaneous tumor incidence of male or female mice. No differences in the incidence of AOM/DSS-induced COAD were observed between these mouse groups, although Ttc22 exon 2&3 deletion partially resulted in the upregulation of adaptive response genes in the colon mucosa. Further study revealed that TPR4 deletion did not disrupt the effect of TTC22 on WTAP upregulation. Consistently, TPR4 deletion did not affect the abundance of total RNA m6A in the colon tissues of the mice, suggesting that the TPR4-deleted Ttc22 mutant remains functional. In conclusion, the TPR4 domain is not essential for the Ttc22 protein. Loss of Ttc22 TPR4 caused no observable changes in the development of C57BL mice or their susceptibility to treatment with the chemical carcinogen AOM/DSS. Whether an essential sequence for a target gene is knocked out should be carefully evaluated before a gene knockout model is employed in formal experiments.