Metabolomic shifts associated with heat stress in coral holobionts

Abstract

Understanding the response of the coral holobiont to environmental change is crucial to inform conservation efforts. The most pressing problem is "coral bleaching," usually precipitated by prolonged thermal stress. We used untargeted, polar metabolite profiling to investigate the physiological response of the coral species Montipora capitata and Pocillopora acuta to heat stress. Our goal was to identify diagnostic markers present early in the bleaching response. From the untargeted UHPLC-MS data, a variety of co-regulated dipeptides were found that have the highest differential accumulation in both species. The structures of four dipeptides were determined and showed differential accumulation in symbiotic and aposymbiotic (alga-free) populations of the sea anemone Aiptasia (Exaiptasia pallida), suggesting the deep evolutionary origins of these dipeptides and their involvement in symbiosis. These and other metabolites may be used as diagnostic markers for thermal stress in wild coral.

Fig. 1. Analysis of Hawaiian stony corals.

(A) Images of M. capitata and P. acuta from Kāne‘ohe Bay, O‘ahu. Photo credit: D. Bhattacharya, Rutgers University. (B) Experimental design. FS refers to field samples. These are the wild coral individuals that were examined at the end of the thermal stress experiment. (C) Color scores for the (right image) ambient- and high-temperature treated coral species M. capitata and P. acuta at the Hawaiʻi Institute of Marine Biology. Ambient-temperature tanks are shown in variations of red, and high-temperature tanks are shown in variations of blue. The color scores represent a proxy for algal symbiont density in coral holobionts with low values indicating bleaching phenotype. The sharp score decrease for P. acuta under ambient tank conditions is explained by the unexpected warming event that occurred in Kāneʻohe Bay, Oʻahu from which the culture water was drawn. Vertical gray lines indicate sampling points T1 (22 May 2019), T3 (3 June 2019), and T5 (7 June 2019). (D) Structures of MAs identified in the coral holobiont. (E) Accumulation of total MAs in M. capitata over the duration of the tank experiments and from wild populations. (F) Metabolite arginine-glutamine (RQ) in M. capitata changes over time during thermal stress (T1 to T5). (G) The metabolite C₁₁H₂₂N₆O₄ that showed accumulation under heat stress matches the synthetic standard of RQ dipeptide in retention time and MS² spectra.

Fig. 2. Dipeptide production by stony corals under thermal stress.

(A to C) Volcano plots of M. capitata metabolites generated from positive ionization mode data when comparing ambient- and high-temperature treatments. Putative dipeptides are shown with the filled orange circles. RA at T3 had an adjusted P = 1 and is not marked on the plot. (D to F) Accumulation of dipeptides KQ, RV, and RA under heat stress. The P values are all <0.005 when comparing ambient T5 and heat-stressed T5 using Student’s t test. (G to I) The metabolites that showed accumulation under heat stress matched synthetic standards of KQ, RV, and RA dipeptides in retention time and MS² spectra.

Fig. 3. Production of known metabolites by M. capitata under thermal stress.

(A) Accumulation of methionine, (B) accumulation of methionine sulfoxide, and (C) reduction of SAM. Differences between the T5 ambient- and high-temperature groups are significant, as determined by a two-tailed Student’s t test with P ≤ 0.05 for the analyses shown in (A) and (B). SAM showed a downward trend during the treatment but was not significantly different between the ambient- and high-temperature groups.

Fig. 4. Network analysis of M. capitata metabolites.

(A) ADPC scores for ambient and thermal treatments at T1, T3, and T5. (B) Subnetwork at thermal stress T5 showing the relationship between carbohydrate metabolism and osmolyte accumulation. Dipeptides are shown as triangles (under both positive and negative ionization modes), and other metabolites are shown as circles with annotations, when available. Metabolite intensity and type of correlation are shown in the legend. (C) Subnetwork at thermal stress T5 showing the relationship between dipeptide and amino acid accumulation.