Sirtuin-dependent recovery from aerial heat shock: The effects of food ration, thermal history, and sirtuin inhibition on clearance rates and valve gape activity of the California mussel, Mytilus californianus (Conrad)

https://doi.org/10.1016/j.jembe.2020.151510Get rights and content

Highlights

  • Acute heat shock reduced clearance rate in intertidal mussels for 4 high tide cycles.

  • Mussels' recovery from heat shock depended on acclimation ration and thermal history.

  • Sirtuin inhibition altered heat shock recovery in mussels fed a low ration.

  • Sirtuins may play an important role in mussels' recovery from acute heat shock.

Abstract

Intertidal mussels regularly experience seasonal fluctuations in food availability and heat stress over the course of a tidal cycle, yet little is known about the biochemical mechanisms linking food ration and thermal tolerance, despite evidence that food enhances stress tolerance at the organismal level. We hypothesized that sirtuins (food-dependent protein regulators of cellular homeostasis), which may regulate the cellular stress response of mytilids during heat stress, also affect organismal performance during recovery from heat stress. As part of a broader study, we investigated the effects of acclimation food ration, acclimation emersion (air) temperature, and sirtuin inhibitors on the feeding behavior of the California mussel, Mytilus californianus, during recovery from acute aerial heat shock. Mussels were acclimated for 3 wk. to either a low (0.25% algae·g mussel dry wt−1) or high (1.5% algae·g mussel dry wt−1) algal ration and low (20 °C) or high (30 °C) emersion temperatures, in tidal simulator tanks capable of maintaining standard circadian and circatidal rhythms. Following acclimation, mussels were exposed to two sirtuin inhibitors (nicotinamide and suramin) or no inhibitors for one high tide cycle. During the subsequent low tide cycle, all mussels were exposed to an acute aerial heat shock (33 °C) and then allowed to recover in their respective acclimation conditions. Every 12 h during the 48 h before sirtuin inhibition and 48 h following acute heat shock (in the corresponding high tide periods), we recorded clearance rates and valve gape activity. Mussels acclimated to a low food ration significantly reduced clearance rates with heat shock and sirtuin inhibition, but not with heat shock alone, relative to pre-treatment values regardless of emersion temperature. Mussels acclimated to a low food ration and high emersion temperature showed significant reductions in valve gape activity following an acute heat shock following sirtuin inhibition, but not in any of the other acclimation groups. Unexpectedly, heat shock alone reduced clearance rates of mussels acclimated to high food and high emersion temperatures only, without any additional effect by heat shock and sirtuin inhibition. Together, these findings suggest an important role for sirtuins at the organismal level during recovery from heat shock following acclimation to a low food ration.

Introduction

The intertidal zone is a highly variable physical environment and mussels inhabiting the rocky intertidal routinely experience seasonal and daily fluctuations in body temperature, food availability, salinity, and oxygen (Elvin and Gonor, 1979; Morris and Taylor, 1983; Menge, 1992; Helmuth and Hofmann, 2001; Helmuth et al., 2006; Denny et al., 2011; Dowd et al., 2013). The California mussel (Mytilus californianus) is a dominant space holder in rocky intertidal and nearby subtidal communities along the Pacific coast of North America (Paine, 1974), where they form dense aggregations that provide habitat (Suchanek, 1985), thermal refuge (Lathlean et al., 2017), and a source of food (Landenberger, 1968; Garza, 2005) for other intertidal organisms. Mussels also help maintain ecosystem health (Shinen and Morgan, 2009) and contribute to benthic-pelagic coupling (Loo and Rosenbberg, 1996) by filtering the water column at rates of up to 6 l·h−1·mussel−1 (Rao, 1953). Thus, persistence of mussel populations is an integral component of maintaining coastal community structure and function. Climate change models predict that the extreme conditions experienced by intertidal mussels may be exacerbated, including not only due to an increase in the frequency of high degree days (Harley et al., 2006; Morak et al., 2013; Seuront et al., 2019), but also due to a decrease in the availability and quality of nearshore phytoplankton (Winder and Sommer, 2012), which serve as their primary food source. Alterations in the feeding behavior of mussels affects the growth and fitness of individual mussels (Dahlhoff and Menge, 1996; Dahlhoff et al., 2002; Bayne, 2004), which indirectly influences food available for mussel predators and the composition of plankton communities (Norén et al., 1999). Numerous studies have evaluated the effects of environmental variables, such as food availability and water temperature, on feeding rates of marine mussels (for examples, see Schulte, 1975; Riisgård et al., 2006; Saurel et al., 2007), but little is known about controls of filtration and how feeding behavior shifts temporally after stress under different food ration (Maire et al., 2007).

Sensitivity to aerial temperatures during emersion is thought to be the main driver of mussel distributions in the intertidal zone (Harley and Helmuth, 2003) and mussels in the higher intertidal are generally more tolerant to extreme conditions than lower intertidal mussels (Sorte et al., 2019). Increased tolerance seemingly comes at a cost, as high intertidal mussels show decreases in growth, body size (Connor and Robles, 2015), and reproduction, as well as increased mortality relative to their lower intertidal counterparts (Petes et al., 2007, Petes et al., 2008). The stress of living in the intertidal zone may be one factor driving reductions in growth and reproductive output, which may be compounded by reductions in time available for feeding (Blanchette et al., 2007). Under stressful conditions, such as high aerial temperature exposure, mytilids in wild populations reduce their valve gape activity (Kramer et al., 1989; Dowd and Somero, 2013; Gleason et al., 2017; Miller and Dowd, 2017, Miller and Dowd, 2019) during periods of reimmersion. However, other studies have suggested that mussels may not use feeding as a recovery mechanism following emersion (Bayne et al., 1988), leaving the direct relationship between feeding behavior and aerial heat shock unclear. In this study, we specifically investigated whether mussels respond to acute thermal stress by altering their feeding behavior when reimmersed by measuring clearance rates and valve gape activity during multiple high tide periods, before and after an acute heat shock.

This study was undertaken as part of a broader project that aims to understand how previous food ration and thermal history alter the ability of M. californianus to respond to aerial heat stress across multiple levels of biological organization. Both previous food availability and thermal history are known to affect feeding behavior (Schulte, 1975; Riisgård et al., 2003, Riisgård et al., 2006; Saurel et al., 2007; Cheng et al., 2018) and thermal tolerance (Lesser et al., 2010; Schneider et al., 2010; Fitzgerald-Dehoog et al., 2012; Dowd et al., 2013), yet the interactive effects of previous caloric intake and thermal history on feeding behavior and thermal sensitivity are not known and likely complex. For instance, mussels (Schneider et al., 2010; Fitzgerald-Dehoog et al., 2012) and barnacles (Gilman and Rognstad, 2018) fed high algal rations show higher rates of survival and growth when exposed to heat stress than those with a poor diet. Yet, perhaps contradictorily, both high algal abundances (Schulte, 1975; 2013; Riisgård et al., 2006, Riisgård et al., 2013; Pascoe et al., 2009) and acute heat stress (Gleason et al., 2017) are known to reduce feeding and valve gape, respectively. Repeated aerial exposures to elevated temperatures may cause significant reductions in feeding effort, which could negate the benefits of having high food availability. As such, we compared the feeding response to acute heat shock in mussels that had been acclimated to different aerial temperature and algal abundance conditions, following the acclimation conditions used by Fitzgerald-Dehoog et al. (2012). We predicted that mussels that were well-fed and acclimated to higher emersion temperatures would be less affected by an acute heat shock than those fed lower algal concentrations or acclimated to lower aerial temperatures, and would consequently, recover more quickly from the stress (i.e., resume normal feeding behavior).

To investigate the underlying biochemical mechanisms linking food availability and thermal sensitivity at the organismal level (i.e., clearance rate and valve activity), we examined the role of sirtuins, which are important regulators of cellular homeostasis during stress at the level of the proteome (Vasquez and Tomanek, 2019). One of the grand challenges in physiology is linking phenotypic responses across levels of biological organization (Mykles et al., 2010) and often involves searching for cellular and molecular mechanisms that correspond to changes observed at higher levels (i.e., organ or organism). Our previous research suggests that sirtuins (silent information regulator proteins; Loo and Rine, 1995) are involved in the heat shock response and, therefore, thermal sensitivity in mussel congeners (Tomanek and Zuzow, 2010; Vasquez et al., 2017; Vasquez et al., 2020). Sirtuins are protein deacylases, responsible for activating numerous proteins involved in metabolism, oxidative stress, and protein homeostasis (Rodgers et al., 2005; Han et al., 2013) and their activity varies with caloric intake in other species (Houtkeeper et al., 2010; Kapahi et al., 2017; Vasquez and Tomanek, 2019). Thus, we hypothesized that the observed linkage between caloric intake and thermal sensitivity in mussels is regulated by this protein family. To understand the role of sirtuins and their downstream targets in the response to aerial heat stress, we exposed mussels to two sirtuin inhibitors, nicotinamide and suramin (Lawson et al., 2010; Kumar and Lombard, 2018), prior to aerial heat shock exposure in each of our acclimation groups, as previous research in our lab has shown that suramin alters the proteome of mytilds 24 h after application of the inhibitor (Vasquez et al., 2017). Furthermore, recent evidence suggests that sirtuin inhibition reduced feeding behavior and food intake in planarians (Ziman et al., 2020), but it is not clear how food ration and heat stress modify this effect on feeding in mussels. We predicted that inhibition of sirtuins would increase thermal sensitivity in mussels compared to uninhibited mussels, and would therefore, reduce feeding for longer periods of time than the acute heat shock alone.

Section snippets

Mussel collection and experimental design

Adult Mytilus californianus (45–65 mm length) were collected from the intertidal zone at Hazard Canyon Reef, Montano de Oro State Park, Los Osos, CA (35.27° N, 120.90° W) and cleared of all epibionts. Mussels were acclimated for 3 wk. at the California Polytechnic State University Center for Coastal Marine Studies (Avila, CA) in flow-through, ‘tidal-simulator’ tanks that mimicked a 12-h circadian rhythm, semi-diurnal tide, and allowed for control of food ration and water (immersion) and aerial

Results

The combined effects of a) acclimation to different food rations and emersion temperatures, b) acute heat shock, and c) sirtuin inhibition on the clearance rates and valve gape activity of M. californianus are complex. We observed variation in the response of mussels exposed to acute heat shock alone or a heat shock following sirtuin inhibition depending on acclimation ration and emersion temperature conditions.

Discussion

This study was undertaken to evaluate the relationship between previous thermal history, food ration, and sirtuins on the thermal sensitivity of the California mussel, measured as changes in feeding behavior following exposure to acute heat shock during emersion. Global climate models predict both shifts in food availability and the frequency of high temperature events (Harley et al., 2006; Winder and Sommer, 2012) and an adequate understanding of the link between food availability and thermal

Funding

This work was supported by the National Science Foundation to Lars Tomanek (IOS-1557500) and Anne Todgham (IOS-1557496). Additional funding for student support was provided by the Frost Foundation and the Biology Department at California Polytechnic State University. This work was also supported by the California Agricultural Experimental Station of the University of California Davis (CA-D-ASC-2252-H to AET).

Author contributors

LT, AET, MAM, and MCV were responsible for conceptualization, while all authors participated in data curation. MAM did the formal analyses and writing - original draft. All authors contributed to writing - review & editing. LT and AET obtained funding and provided supervision.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

The authors thank the numerous students who assisted with data collection, including O. Carmo, A. Kretschmar, and J. T. Fang. We would also like to thank Dr. A. Shaffner for statistical advice and R. Brewster, D. Brewster, T. Moylan, and J. Felton for their logistical support with designing and maintaining the tidal simulators at the Cal Poly Pier. And, finally, thank you to our reviewers for making helpful suggestions and overall improving the data analyses and manuscript.

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