First published online October 5, 2006
Journal of Experimental Biology 209, 4102-4114 (2006)
Published by The Company of Biologists 2006
doi: 10.1242/jeb.02484
Seasonal acquisition of chill tolerance and restructuring of membrane glycerophospholipids in an overwintering insect: triggering by low temperature, desiccation and diapause progression
A. Tomcala1,2,
M. Tollarová1,2,
J. Overgaard3,
P. Simek1 and
V. Kostál1,2,*
1 Biology Centre AS CR, Institute of Entomology, Branisovská
31, 370 05 Ceské Budejovice, Czech
Republic
2 Faculty of Biological Sciences, University of South Bohemia, Ceské
Bud
jovice, Czech Republic
3 Department of Terrestrial Ecology, National Environmental Research
Institute, Silkeborg, Denmark
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Fig. 1. Course of air temperatures in the field (A) and the temperature protocol
used for laboratory acclimation (B) of Pyrrhocoris apterus. The adult
insects were sampled on 23 October and 27 December in the field. Laboratory
adults were maintained under short days conditions (12 h:12 h, L:D). Their
physiological state was considered as the initial state when they reached age
of 4 weeks. Three different treatments were then applied: progression of
diapause, desiccation, and cold acclimation (see text for detailed
descriptions).
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Fig. 2. Electrospray ionisation mass spectra showing molecular species composition
of P. apeterus thoracic muscle tissue. Positive full mass spectra of
(A) glycerophosphoethanolamine (GPEtn) and (B) glycerophosphocholine (GPChol)
fractions are depicted separately. Identities of all major [M+H]+
molecular ions (solid ovals) were verified by MS2 and MS3 fragmentations and
are shown in Fig. 3. Note that
each major ion occurs as a pair of masses differing by 1 amu (13C
isotopic contribution). All species formed the adducts with sodium
[M+Na]+ in relatively low and constant proportions (broken
ovals).
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Fig. 3. HPLC separation of major molecular species of (A)
glycerophosphoethanolamines (GPEtns) and (B) glycerophosphocholines (GPChols)
of P. apeterus thoracic muscle tissue. The upper most trace (in A and
B) shows total GPEtn and GPChol fractions and the lower traces show individual
ion masses (molecules), as they were extracted using the MS ion trap. Note
that a relative y axis are used in all chromatograms, where the
highest peak is taken as 100% response. That is why the signal/noise is
relatively low in the rare species, e.g. 16:0/18:1-GPChol. For quantification
purposes, the areas under chromatographic peaks of individual molecular
species derived from a total chromatogram (upper traces) were compared.
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Fig. 4. Restructuring of glycerophospholipid composition in thoracic muscle
membranes of Pyrrhocoris apterus during field acclimatization between
23 October and 27 December. (A) Mean (± s.d.; N=6 independent
samples) relative proportion of phosphatidylethanolamine (GPEtn) and
phosphatidylcholine (GPChol) molecule (all species make 100% in total) in
October. (B) The difference from the initial state at the end of the
acclimatization period. The differences were statistically analyzed using
unpaired two-tailed t-tests (*P<0.05;
**P<0.001; ***P<0.0001).
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Fig. 5. Restructuring of glycerophospholipid composition in fat body tissue
membranes of Pyrrhocoris apterus during the field acclimatization
between 23 October and 27 December. (A) Mean (± s.d.; N=6
independent samples) relative proportion of phosphatidylethanolamine (GPEtn)
and phosphatidylcholine (GPChol) molecule (all species make 100% in total) in
October. (B) The difference from the initial state at the end of the
acclimatization period. The differences were statistically analyzed using
unpaired two-tailed t-tests (*P<0.05;
**P<0.001; ***P<0.0001).
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Fig. 6. Restructuring of glycerophospholipid composition in thoracic muscle
membranes of Pyrrhocoris apterus after various laboratory
acclimations. (A) Initial state; (B) dipause progression; (C) desiccation; (D)
cold acclimation. The differences were statistically analyzed using unpaired
two-tailed t-tests (*P<0.05; **P<0.001;
***P<0.0001).
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Fig. 7. Restructuring of glycerophospholipid composition in fat body tissue
membranes of Pyrrhocoris apterus after various laboratory
acclimations. (A) Initial state; (B) dipause progression; (C) desiccation; (D)
cold acclimation. The differences were statistically analyzed using unpaired
two-tailed t-tests (*P<0.05; **P<0.001;
***P<0.0001).
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Fig. 8. Results of multivariate principal component analysis of the differences
between membrane glycerophospholipid compositions in thoracic muscles and fat
body tissues of Pyrrhocoris apterus sampled in the field (23 October
and 27 December) or variously acclimated in the laboratory (initial state,
diapause progression, desiccation, cold acclimation). (A) The centres of
elipses represent the means, and the borders the standard errors, of the
scores from the first and second principal component. The dashed line
separates the data for muscles and fat-body tissues. (B,C) The differences
among the scores of PC1 and PC2, respectively, were tested using one-way ANOVA
followed by a Student–Newman–Keuls test (different letters
indicate statistically different values).
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© The Company of Biologists Ltd 2006
