Expression Patterns of the Heat Shock Protein 90 (Hsp90) Gene Suggest Its Possible Involvement in Maintaining the Dormancy of Dinoflagellate Resting Cysts

Abstract

Heat shock protein 90 (Hsp90) is a highly conserved molecular chaperone functioning in cellular structural folding and conformational integrity maintenance and thus plays vital roles in a variety of biological processes. However, many aspects of these functions and processes remain to be fully elucidated, particularly for non-model organisms. Dinoflagellates are a group of eukaryotes that are exceedingly important in primary production and are responsible for the most harmful algal blooms (HABs) in aquatic ecosystems. The success of dinoflagellates in dominating the plankton community is undoubtedly pertinent to their remarkable adaptive strategies, characteristic of resting cyst production and broad tolerance to stresses of temperature and others. Therefore, this study was conducted to examine the putative roles of Hsp90 in the acclimation to temperature stress and life stage alterations of dinoflagellates. Firstly, we isolated the full-length cDNA of an Hsp90 gene (StHsp90) via RACE from the cosmopolitan HAB species Scrippsiella trochoidea and tracked its transcriptions in response to varied scenarios via real-time qPCR. The results indicated that StHsp90 displayed significant mRNA augment patterns, escalating during 180-min treatments, when the cells were exposed to elevated and lowered temperatures. Secondly, we observed prominently elevated StHsp90 transcriptions in the cysts that were stored at the cold and dark conditions compared to those in newly formed resting cysts and vegetative cells. Finally, and perhaps most importantly, we identified 29 entries of Hsp90-encoding genes with complete coding regions from a dinoflagellate-specific environmental cDNA library generated from marine sediment assemblages. The observed active transcription of these genes in sediment-buried resting cysts was fully supported by the qPCR results for the cold-stored resting cysts of S. trochoidea. Hsp90s expressions in both laboratory-raised and field-collected cysts collectively highlighted the possible involvement and engagement of Hsp90 chaperones in the resting stage persistence of dinoflagellates.

Keywords: Scrippsiella trochoidea; dinoflagellate; dormancy; environmental cDNA library; heat shock protein 90 (Hsp90); resting cysts; temperature stress.

Figure 1

The transcription levels of StHsp90 in cells under different shock temperatures (A) and during a 180-min time course with a 30 °C treatment (B) and 10 °C exposure (C). Each bar represents the mean of three biological replicates, and error bars signify the standard deviation of those means. Asterisk indicates significant increases at the transcription level as compared with the 20 °C group (p < 0.05) (A). Same letter above the bar denotes no significant difference (p > 0.05) in abundance (B,C).

Figure 2

The transcription levels of StHsp90 in cells exposed to one-step or stepwise temperature shock (+10 °C (A) and −10 °C (B)). One-step shock: put the cells directly in a temperature variation of 10 °C for 60 min. Stepwise shock: first, put the cells in a temperature variation of 5 °C for 10 min and then subject them to a further 5 °C change for 60 min. The 20 °C maintained cultures are used as the control. The relative expression values are shown as the mean fold changes compared to the control, with the error bars depicting the standard deviations (n = 3). Same letter above the bars denotes no significant difference (p > 0.05) in abundance.

Figure 3

The transcription levels of StHsp90 in cells at different life stages (vegetative cells at the exponential and stationary growth stages, newly formed resting cysts, and resting cysts in a forced dormancy at a lower temperature and in darkness for 20 and 30 days, respectively). The relative expression values are shown as the mean fold changes compared to the control (vegetative cells collected at Day 5), with the error bars depicting the standard deviations (n = 3). Significant differences are indicated with different letters at p < 0.05.

Figure 4

The Bayesian inference (BI) distance tree formulated from the alignment of the full amino acid sequences of the complete coding regions of the 90 Hsp90 sequences. Numbers at the nodes represent the BI posterior probabilities. For the 60 entries downloaded from the GenBank database, the accession number of each sequence is noted following the species name (see Supplementary Table S1 for more details). The newly generated Hsp90 gene from Scrippsiella trochoidea in the current study is boxed. The 29 newly identified Hsp90 sequences from the e-cDNA library are present as the GenBank accession number highlighted in the gray background.