Hormonal control of hemolymph lipoprotein ice nucleators in overwintering freeze-susceptible larvae of the stag beetle Ceruchus piceus: adipokinetic hormone and juvenile hormone

Summary Freeze-resistant overwintering larvae of the stag beetle Ceruchus piceus do not produce antifreezes in winter, but instead lower their supercooling points by seasonal removal of lipoprotein ice nucleators (LPINs) from the hemolymph. The normal lipid transport function of these lipoproteins becomes less essential during winter because of the low temperatures and the diapause state of the larvae. Adipokinetic hormone (AKH) and juvenile hormone (JH) were shown to be involved in the control of supercooling abilities and LPIN levels. Treatment of midwinter larvae with AKH resulted in an increase in ice nucleator activity within 2 h, associated with elevated levels of LPINs, as demonstrated by Western blots derived from native PAGE gels probed with polyclonal antibodies to the LPINs. AKH also stimulated the release of LPIN in vitro from cultured fat bodies. In contrast, JH treatments of larvae with high hemolymph ice nucleator contents (either autumn or spring larvae) caused a decrease in ice nucleator activity and supercooling points. However, Western blots showed increased LPIN levels in these JH treated larvae. Apparently, this JH-induced, inactive form of LPIN lacks some component(s) essential for ice nucleator activity.Abbreviations AKH Adipokinetic hormone - Apo-I Apolipoprotein-I - Apo-II Apolipoprotein-II - JH Juvenile hormone - LPIN Lipoprotein ice nucleator - PAGE Polyacrylamide gel electrophoresis - SDS Sodium dodecyl sulfate - SCP Supercooling point - THP Thermal hysteresis protein - HDLp High density lipophorin - VHDLp Very high density lipophorin