Intracellular Localization and Gene Expression Analysis Provides New Insights on LEA Proteins’ Diversity in Anhydrobiotic Cell Line

SIMPLE SUMMARY: Polypedilum vanderplanki (sleeping chironomid) is widely known for its ability to withstand complete desiccation in a state of anhydrobiosis. The genome of this insect contains a number of hugely expanded paralogous gene groups, including 27 genes that encode late embryogenesis abundant (LEA) proteins. An important question regarding such paralogous genes is whether they are functionally specialized or not. Previously, we found that PvLEA proteins in C-terminal fusions with green fluorescent protein (AcGFP1) have four distinct localization types in mammalian cells. In the current paper, we studied PvLEA expression and localization in both N- and C-terminal fusions with AcGFP1 in anhydrobiotic Pv11 cells, derived from P. vanderplanki. We found that all but two PvLea genes are expressed in Pv11 cells and are upregulated during anhydrobiosis-inducing trehalose treatment similarly to the larvae of P. vanderplanki during the real induction of anhydrobiosis. We found that the localization of PvLEA proteins in N-terminal fusions with AcGFP1 is highly uniform in Pv11 cells and the Sf9 insect cell line. We observed an inconsistency of PvLEA localization between different cell cultures and between N- and C-terminal fusions, that needs to be taken into account when using PvLEA in the engineering of anhydrobiotic cell lines. ABSTRACT: Anhydrobiosis, an adaptive ability to withstand complete desiccation, in the nonbiting midge Polypedilum vanderplanki, is associated with the emergence of new multimember gene families, including a group of 27 genes of late embryogenesis abundant (LEA) proteins (PvLea). To obtain new insights into the possible functional specialization of these genes, we investigated the expression and localization of PvLea genes in a P. vanderplanki-derived cell line (Pv11), capable of anhydrobiosis. We confirmed that all but two PvLea genes identified in the genome of P. vanderplanki are expressed in Pv11 cells. Moreover, PvLea genes are induced in Pv11 cells in response to anhydrobiosis-inducing trehalose treatment in a manner highly similar to the larvae of P. vanderplanki during the real induction of anhydrobiosis. Then, we expanded our previous data on PvLEA proteins localization in mammalian cells that were obtained using C-terminal fusions of PvLEA proteins and green fluorescent protein (GFP). We investigated PvLEA localization using N- and C-terminal fusions with GFP in Pv11 cells and the Sf9 insect cell line. We observed an inconsistency of PvLEA localization between different fusion types and different cell cultures, that needs to be taken into account when using PvLEA in the engineering of anhydrobiotic cell lines.