Deciphering the phosphorylation-based regulatory strategies of Haemaphysalis longicornis in heat stress.

Background: The Asian hard tick (Haemaphysalis longicornis) is an obligate hematophagous ectoparasite belonging to the family Ixodidae (hard ticks). This species serves as a competent vector for numerous highly pathogenic agents. The number of ticks that survive the summer, particularly under high-temperature conditions, exerts immediate and lasting impacts on tick populations and tick-borne disease prevalence. Therefore, it is crucial to study how high temperatures affect ticks, as well as how ticks adopt effective behavioral strategies and physiological adaptations to cope with heat stress. Phosphorylation, a kind of important protein post-translational modification (PTM), is vital for cellular signal transduction, gene expression, and rapid cell cycle regulation.

Methods: This study systematically analyzed phosphorylation changes in proteins from the salivary gland, midgut, ovary, and Malpighian tubules of ticks exposed to different temperatures (26 °C, 36 °C, and 45 °C) using quantitative proteomics. Differentially expressed phosphoproteins were comprehensively assessed using bioinformatics tools, supplemented with ribonucleic acid (RNA) interference and tick survival assays to validate key protein functions.

Results: This study reveals a tissue-specific phosphorylation regulatory pattern. It identifies the involvement of kinase families such as CK1, AGC, and CMGC in the heat stress response. Phosphorylation modifications of spliceosome components and upregulated Hsp90 phosphorylation were found to regulate RNA splicing pathway and heat shock response, respectively. Notably, the Hsp90 co-chaperone CDC37 was critical for maintaining GRK stability and ensuring tick survival under high-temperature conditions.

Conclusions: The thermal stress response in H. longicornis involves a coordinated network of protein kinases, alternative splicing events, and heat shock proteins along with their co-chaperones. These findings provide a foundation for further deciphering of the molecular regulatory mechanisms of tick tolerance to high temperatures.