Nucleoplasmin‐Like Protein, Expression Differences During the Caste Polyphenism Processes in Apis mellifera

ABSTRACT

In honey bees, queen–worker differentiation is regulated by diet and begins after the first 3 days post‐hatching. Several molecules and processes appear to be involved in caste polyphenism, including juvenile hormone, royalactin, and DNA methylation. A protease (PS4) was recently detected in worker larvae at 72 h post‐hatching and shown to be inhibited by royal jelly, suggesting a potential role in this process. This study investigated the expression and tissue localization of nucleoplasmin during caste polyphenism in Apis mellifera larvae. A protein spot corresponding to a previously described caste‐specific proteolytic activity was separated by 2DE and identified by mass spectrometry as a nucleoplasmin‐like protein. Based on this result, a polyclonal antibody was raised against a conserved nucleoplasmin peptide and subsequently used for immunoproteomic and immunohistochemical analyses in worker‐ and queen‐destined larvae. At this instar, Western blot analysis using a polyclonal anti–nucleoplasmin‐like protein antibody revealed bands between 43–88 kDa and 74–104 kDa in worker‐ and queen‐destined larvae, respectively, which may correspond to different polymeric conformations of the nucleoplasmin‐like protein. Differences in the molecular mass of nucleoplasmin‐like protein spots between worker and queen larvae suggest caste‐specific roles during differentiation. Immunohistochemistry showed a ubiquitous nuclear distribution of the protein in undifferentiated larvae. Worker‐fate larvae diverge from this pattern as early as Day 4, whereas queen‐fate larvae retain a distribution like that of undifferentiated larvae, possibly representing the baseline developmental condition. Further studies examining temporal changes in nucleoplasmin‐like protein expression between the third and fourth larval instars and during post‐larval stages are desirable.