Heat-shock proteins
   
 

A considerable literature has developed around the subject of heat-shock proteins (HSPs) in intertidal marine invertebrates. These are special molecules known as molecular chaperones that function to ensure proper 3-dimensional folding and compartmentalisation of proteins.  After heat stress HSPs prevent aggregation of heat-damaged proteins and facilitate re-naturation of a protein’s tertiary structure.  An individual snail (as an example) may have several different types of HSPs, each being expressed at a different temperature.  Considered here are papers on HSPs in black turban snails Chlorostoma (Tegula) funebralis.

The topic of heat-shock proteins is considered here, while other topics including energy imbalances in veliger larvae of abalones and microhabitat effects on internal salt concentrations in black turban snails are considered in another general section on PHYSIOLOGICAL ECOLOGY.

HSPs in other west-coast invertebrates can be found elsewhere in the ODYSSEY: LEARN ABOUT MUSSELS: LIFE IN THE INTERTIDAL ZONE: HEAT-SHOCK PROTEINS and LEARN ABOUT LIMPETS: LIFE IN THE INTERTIDAL ZONE: TEMPERATURE STRESS

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Research study 1
 

photographs of several trochid snails including Chlorostoma funebralis with indication of preferred tidal heights inhabitedgraph showing body temperatures of trochid snails Chlorostoma funebralis and Chlorostoma brunnea during springtime at Pacific Grove, CaliforniaIn addition to desiccation stresses in their intertidal habitat, Chlorostoma funebralis are subject to seasonal thermal stresses.  At Pacific Grove, California only the warm seasonal stresses are important, but further north winter freezing may also be a critical limiting factor.  Three Chlorostoma species co-occur at Pacific Grove:  graph showing survival of trochid snails Chlorostoma funebralis and C. brunnea at different temperaturesfunebralis in the mid-low intertidal region where body temperatures regularly exceed 33oC during spring and summer, and brunnea and  montereyi in the low intertidal-subtidal region where body temperatures rarely exceed 20-25oC. As the graph above Right shows, not only are body1 temperatures overall higher for funebralis as compared with the subtidal-inhabiting brunnea, but the rate of temperature change is much greater in the higher-intertidal habitat, especially during spring tides. 

Heat tolerances would be expected to differ among the 3 species of Chlorostoma.  This is tested in at Hopkins Marine Station at Pacific Grove, California by subjecting 20 individuals of each species to temperature increases of 1OoC every 12min and measuring percentage survival.  Not surprisingly, the 2 subtidal species die at much lower temperatures than the intertidal species.  LT502 (=LD50) values are 36oC for C. brunnea and C.montereyi, and 43oC for C. funebralis (see graph lower Right).

graphs showing relative levels of HSPs in trochid snails at different temperaturesInvestigation of heat-shock proteins (HSPs) in these 3 Chlorostoma species shows3, as one would predict, that induction temperature is higher in funebralis (27oC) than in brunnea or montereyi (=24oC; see graphs lower Left).  In fact, the two subtidal species rarely experience temperatures that would result in enhanced expression of HSPs.  For example, during the period shown in the display (above Right) there is only 1d where 24oC body temperature is exceeded in brunnea, whereas there are 11d  where 27oC body temperature is exceeded in funebralis. The HSP response is energetically costly, both in the synthesis of the HSPs and in their chaperoning activity.  In this regard, then, funebralis likely incurs greater costs than the other two species.  The authors note that upper thermal limits for synthesis of HSPs differ in the 3 species.  In fact, some protein synthesis in the two subtidal species is heat-inactivated at temperatures commonly experienced by funebralis. This alone could prevent these species from inhabiting the mid-low intertidal region occupied by funebralis.  Tomanek & Somero 1999 J Exper Biol 202: 2925; for a brief review of thermal stress and HSPs in Chlorostoma and other marine invertebrates see Tomanek & Helmuth 2002 Integ Comp Biol 42: 771.

NOTE1  the data presented are actually from gelatin-filled shells of each species carrying implanted thermistor probes.  Comparisons with living snails show that the mimic “tissue” has similar thermal responses to that of living tissue

NOTE2 lethal temperature-50 (LT50) or lethal death-50 (=LD50), is the temperature at which 50% of the test animals are dead

NOTE3 the graphs show levels of hsp70 in the 3 species at 13oC (field seawater temperature) and then at a "stress" temperature of 18oC (other temperature profiles are provided by the authors but not shown here)

 

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Research study 2

 

graph showing survival of trochid snails Chlorostoma funebralis and C. brunnea over 50h at 30 degrees C Further research on Chlorostoma spp. at Hopkiins Marine Station, Pacific Grove, California investigates the time course and magnitude of heat-shock response in C. funebralis, a mainly intertidal species, and C. brunnea, a low intertidal/subtidal species.  The authors subject test groups of each species to 2.5h immersion in 30oC seawater, transfer them to 13oC and monitor survival and levels of several HSPs over an additional 50h.  This level of thermal stress is not extreme for funebralis, which commonly experiences summer body temperatures of 30oC, but is extreme for the deeper-living brunnea which rarely is air-exposed.  In fact, the 30oC temperature stress is eventually lethal to all brunnea after about 50h.

graph showing levels of HSPs in trochid snails Chlorostoma brunnea and C. funebralis at different temperaturesWithin 2h following the heat stress both species begin to produce HSP70, but levels fall off to initial levels within about 6h for funebralis.  In contrast, levels continue to histogram showing relative levels of HSPs in trochid snials Chlorostoma funebralis and C. brunnea in air and water at 30 degrees Cbuild in brunnea, but later decrease somewhat, presumably in response to the onset of death. 

Interestingly, if the 30oC heat shock is applied to air-exposed animals, then the HSP-synthesis response is greatly magnified in brunnea but not in funebralis. The data suggest that funebralis may be able to repair protein damage incurred during one mid-day low tide before the occurrence of the next low tide.  Moreover, sufficient heat-shock proteins may accumulate from one heat stress to cope with the next bout of heat stress.  The different time courses and magnitudes of synthesis of heat-shock proteins in the 2 species support the idea that vertical limits of distribution may in part attribute to thermal stress.  Tomanek & Somero 2000 Physiol Biochem Zool 73: 249; see also Tomanek 2002 Integ Comp Biol 42: 797 and Tomaneck & Somero 2002 J Exp Biol 205: 677.


NOTE  the authors present data for several HSPs, but only those for HSP 70 are shown here

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Research study 3
 

graphs showing effect of different field conditions on levels of heat-shock proteins HSP72 in snails Chlorostoma funebralis and C. brunneaHow variable over time are HSPs, and how much do they vary with intertidal position and under changing environmental conditions?  These questions are addressed using the same Chlorostoma species as used in Research Study 2 above, also at Hopkins Marine Station, Pacific Grove, California.  Specimens of C. brunnea are translocated from their usual low-intertidal/subtidal positions to shaded and unshaded cages at the mid-intertidal level to subject them to increased stress.  Specimens of C. funebralis are also translocated but, in this case of these already intertidal individuals, from parts of the shore normally occupied to shaded and unshaded mid-intertidal cages.  Snails in cages and in their original populations are sampled every 3-4d for 1mo, and endogenous levels of HSP72 and HSP74 quantified.  In situ body temperatures are recorded by data-loggers.

Results show that after periods of mid-day low tides, HSP72 levels increase more in caged C. brunnea than in caged C. funebralis (see upper graph).Levels in C. brunnea increase more in sun-exposed cages than in shaded ones, suggesting that greater emersion and reduced feeding time are actually less stressful than exposure to sun. Exposure to sun in C. funebralis has a significant effect over normal field-acclimatised individuals, but only under certain stressful combinations of tide and temperature (see lower graph). The results support the authors’ hypothesis that Chlorostoma spp. exposed to greater temperature extremes and fluctuations at higher intertidal levels will activate their heat-shock responses accordingly.  Tomanek & Sanford 2003 Biol Bull 205: 276; for a good review of thermal physiology in relation to intertidal zonation in gastropods, crabs, and other animals see Somero 2002 Integr Comp Biol 42: 780.


NOTE  the classification of HSPs is based on their molecular mass in kDa.  Only results for HSP72 are shown here; those for HSP74 are generally similar 

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