Abstract: Objective To investigate the differences between two well recognized WiskottAldrich syndrome protein and SCAR homolog ( WASH) with different sequences. Methods Immunofluorescence, immunoprecipitation, and laser micro-irradiation were used to analyze the differences between WASH468 and WASH465 in localization patterns, the interaction with FAM21 or Ku proteins, the recruitment to DNA damage sites and the protein stabilities. To analyze the universality and conservation of the two types of WASH sequences, the WASH protein sequences in various organisms were compared, and the WASH coding sequences in various cell types commonly used forbiological and medical research were detected. Results Both WASH exhibited a similar way in localizing to endosome. WASH468 exhibited a stronger interaction with FAM21, and was more stable than WASH465. WASH465 exhibited a stronger interaction with Ku protein, and the recruitment of WASH465 to DNA damage sites was more robust than that of WASH468. The degree of similarity observed between the sequence of WASH468 in humans and the sequences of WASH in other organisms was significantly greater than that observed with WASH465. Moreover, the coding sequences of WASH in various cell types, which are commonly utilized in biological and medical research, exhibited concordance with the sequence of WASH468. Conclusion There are differences in the biological characteristics between WASH468 and WASH465, and WASH468 is significantly more universal and conserved than WASH465. Therefore, WASH468 is the conserved human WASH sequence.

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