The IRE1 pathway regulates honey bee Unfolded Protein Response gene expression.

Our molecular understanding of honey bee cellular stress responses is incomplete. Previously, we sought to identify and began functional characterization of the components of the Unfolded Protein Response (UPR) in honey bees. We observed that UPR stimulation resulted in induction of target genes upon IRE1 pathway activation, as assessed by splicing of Xbp1 mRNA. However, we were not able to determine the relative role of the various UPR pathways in gene activation. Our understanding of honey bee signal transduction and transcriptional regulation has been hampered by a lack of tools. After using RNA-seq to expand the known UPR targets in the honey bee, we used the Drosophila melanogaster S2 cell line and honey bee trans and cis elements to investigate the role of the IRE1 pathway in the transcriptional activation of one of these targets, the honey bee Hsc70-3 gene. Using a luciferase reporter, we show that honey bee Hsc70 promoter activity is inducible by UPR activation. In addition, we show that this activation is IRE1-dependent and relies on specific cis regulatory elements. Experiments using exogenous honey bee or fruit fly XBP1S proteins demonstrate that both factors can activate the Hsc70-3 promoter and further support a role for the IRE1 pathway in control of Hsc70-3 expression in the honey bee. By providing foundational knowledge about the UPR in the honey bee and demonstrating the usefulness of a heterologous cell line for molecular characterization of honey bee pathways, this work stands to improve our understanding of this critical species.

The heat shock response and humoral immune response are mutually antagonistic in honey bees.

The honey bee is of paramount importance to humans in both agricultural and ecological settings. Honey bee colonies have suffered from increased attrition in recent years, stemming from complex interacting stresses. Defining common cellular stress responses elicited by these stressors represents a key step in understanding potential synergies. The proteostasis network is a highly conserved network of cellular stress responses involved in maintaining the homeostasis of protein production and function. Here, we have characterized the Heat Shock Response (HSR), one branch of this network, and found that its core components are conserved. In addition, exposing bees to elevated temperatures normally encountered by honey bees during typical activities results in robust HSR induction with increased expression of specific heat shock proteins that was variable across tissues. Surprisingly, we found that heat shock represses multiple immune genes in the abdomen and additionally showed that wounding the cuticle of the abdomen results in decreased expression of multiple HSR genes in proximal and distal tissues. This mutually antagonistic relationship between the HSR and immune activation is unique among invertebrates studied to date and may promote understanding of potential synergistic effects of disparate stresses in this critical pollinator and social insects more broadly.

Divergent forms of endoplasmic reticulum stress trigger a robust unfolded protein response in honey bees.

Honey bee colonies in the United States have suffered from an increased rate of die-off in recent years, stemming from a complex set of interacting stresses that remain poorly described. While we have some understanding of the physiological stress responses in the honey bee, our molecular understanding of honey bee cellular stress responses is incomplete. Thus, we sought to identify and began functional characterization of the components of the UPR in honey bees. The IRE1-dependent splicing of the mRNA for the transcription factor Xbp1, leading to translation of an isoform with more transactivation potential, represents the most conserved of the UPR pathways. Honey bees and other Apoidea possess unique features in the Xbp1 mRNA splice site, which we reasoned could have functional consequences for the IRE1 pathway. However, we find robust induction of target genes upon UPR stimulation. In addition, the IRE1 pathway activation, as assessed by splicing of Xbp1 mRNA upon UPR, is conserved. By providing foundational knowledge about the UPR in the honey bee and the relative sensitivity of this species to divergent stresses, this work stands to improve our understanding of the mechanistic underpinnings of honey bee health and disease.