Post-diapause synthesis of ArHsp40-2, a type 2 J-domain protein from Artemia franciscana, is developmentally regulated and induced by stress.

Nathan M Rowarth,Thomas H Macrae,Jyotshna Kanungo

doi:10.1371/journal.pone.0201477

Abstract

Post-diapause cysts of Artemia franciscana undergo a well-defined developmental process whereby internal differentiation leads to rupture of the cyst shell, release of membrane-enclosed nauplii and hatching to yield swimming larvae. The post-diapause development of A. franciscana has been examined at biochemical and molecular levels, yet little is known about molecular chaperone function during this process. In addressing this we recently described ArHsp40, a type 1 J-domain protein in post-diapause A. franciscana cysts and larvae. The current report describes ArHsp40-2, a second J-domain protein from A. franciscana. ArHsp40-2 is a type 2 J-domain protein, lacking a zinc binding domain but containing other domains characteristic of these proteins. Notably, ArHsp40-2 possesses a double barrel β-domain structure in its substrate binding region, as does ArHsp40. qPCR revealed a relatively low amount of ArHsp40-2 mRNA in 0 h cysts which increased significantly until the E1 stage, most likely as a result of enhanced transcription, after which it declined. An antibody specific to ArHsp40-2 was produced and used to show that like its mRNA, ArHsp40-2 accumulated until the E1 stage and then decreased to amounts lower than those in 0 h cysts. The synthesis of ArHsp40-2 was induced by heat shock indicating that ArHsp40-2 is involved in stress resistance in cysts and nauplii. Accumulation in cysts during early post-diapause development followed by its sharp decline suggests a role in protein disaggregation/refolding, a function of Hsp40s from other organisms, where ArHsp40-2 assists in the rescue of proteins sequestered during diapause by p26, an abundant small heat shock protein (sHsp) in A. franciscana cysts.

Highlights

Fertilized eggs of the extremophile crustacean, Artemia franciscana develop ovoviviparously in the female’s brood sac and at five days post-fertilization swimming nauplii are released [1, 2]
The carboxyl-terminal domain contained a double barrel β-domain followed by an α-helix which forms a dimerization domain similar to that found in ArHsp40 and other J-domain proteins (Fig 5)
RNA interference (RNAi) experiments confirm that p26 has an important role in protecting diapause cysts during stress [9], but because this small heat shock protein (sHsp) lacks foldase activity other molecular chaperones including Hsp110, Hsp90 and Hsp70, and their accessory proteins such as the Hsp40s, are required to refold proteins

Summary

Introduction

Fertilized eggs of the extremophile crustacean, Artemia franciscana develop ovoviviparously in the female’s brood sac and at five days post-fertilization swimming nauplii (larvae) are released [1, 2]. Under optimal conditions larvae molt several times and reach sexual maturity in four to five weeks. With changing parameters such as photoperiod and temperature, oviparous. The encysted embryos, termed cysts, are released from the brood sac and after six to eight days enter diapause, a physiological state of reduced metabolism and enhanced stress tolerance that allows survival under adverse environmental conditions [4,5,6,7]. Once diapause terminates the encysted embryos resume growth if conditions are favourable and the resulting nauplii develop in the same way as nauplii arising ovoviviparously

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Conclusion