Sex-specific transgenerational plasticity II: Grandpaternal effects are lineage- and sex-specific in threespined sticklebacks

1. Transgenerational plasticity (TGP) occurs when the environment encountered by one generation (F0) alters the phenotypes of one or more future generations (e.g. F1 and F2). Sex selective TGP, via specific lineages or to only male or female descendants, has been underexplored in natural systems, and may be adaptive if it allows past generations to fine-tune the phenotypes of future generations in response to sex-specific life history strategies. 

2. We sought to understand if exposing males to predation risk can influence grandoffspring via sperm in threespined stickleback (Gasterosteus aculeatus). We specifically tested the hypothesis that grandparental effects are transmitted in a sex-specific way down the male lineage, from paternal grandfathers to F2 males.  

3. We reared F1 offspring of unexposed and predator-exposed F0 males under ‘control’ conditions and used them to generate F2s with control grandfathers, a predator-exposed maternal grandfather (i.e., predator-exposed F0 males to F1 daughters to F2 offspring), a predator-exposed paternal grandfather (i.e., predator-exposed F0 males to F1 sons to F2 offspring), or two predator-exposed grandfathers. We then assayed male and female F2s for a variety of traits related to antipredator defense. 

4. We found little evidence that transgenerational effects were mediated to only male descendants via the paternal lineage. Instead, grandpaternal effects depended on lineage and were mediated largely across sexes, from F1 males to F2 females and from F1 females to F2 males. When their paternal grandfather was exposed to predation risk, female F2s were heavier and showed a reduced change in behavior in response to a simulated predator attack relative to offspring of control, unexposed grandparents. In contrast, male F2s showed reduced antipredator behavior when their maternal grandfather was exposed to predation risk. However, these patterns were only evident when one grandfather, but not both grandfathers, was exposed to predation risk, suggesting the potential for non-additive interactions across lineages.

5. If sex-specific and lineage effects are common, then grandparental effects are likely underestimated in the literature. These results draw attention to the importance of sex-selective inheritance of environmental effects and raise new questions about the proximate and ultimate causes of selective transmission across generations.

Methods
All behavioral and morphological data were collected live on paper and transcribed electronically. Cortisol data were calculated from OD scores.

1. 代际可塑性（Transgenerational plasticity, TGP）指某一世代（F0）所经历的环境，会改变一个或多个后续世代（如F1与F2）的表型。通过特定谱系、或仅影响雄性/雌性后代的性别选择性代际可塑性，在自然系统中尚未得到充分探索；若其能让祖先世代依据性别特异的生活史策略微调后代表型，则该机制可能具有适应性。

2. 本研究旨在探究：在三棘刺鱼（Gasterosteus aculeatus）中，使雄性暴露于捕食风险能否通过精子对隔代表型产生影响。我们专门检验了如下假说：祖代效应以性别特异的方式沿雄性谱系传递，即从父系祖父传递至F2雄性个体。

3. 我们在“对照”条件下饲养未暴露与暴露于捕食风险的F0雄性的F1后代，并利用这些F1个体培育得到四类F2个体：拥有对照祖父的F2、拥有捕食风险暴露的母系祖父的F2（即：捕食风险暴露的F0雄性→F1雌性→F2后代）、拥有捕食风险暴露的父系祖父的F2（即：捕食风险暴露的F0雄性→F1雄性→F2后代），以及祖父双方均暴露于捕食风险的F2。随后，我们对F2雌雄个体的多项反捕食防御相关性状进行了测定。

4. 研究结果显示，几乎没有证据表明代际效应仅通过父系谱系传递给雄性后代。相反，祖代效应依赖于谱系，且主要以跨性别方式介导：从F1雄性传递至F2雌性，以及从F1雌性传递至F2雄性。当父系祖父暴露于捕食风险时，F2雌性个体体重更大，且在模拟捕食攻击下的行为变化幅度相较于未暴露祖父的对照后代有所降低。与之相对，当母系祖父暴露于捕食风险时，F2雄性个体的反捕食行为有所减弱。不过，这些模式仅在仅一位祖父（而非两位祖父）暴露于捕食风险时才会显现，这提示谱系间存在非加性交互作用的可能性。

5. 若性别特异且谱系依赖的效应普遍存在，则现有文献中对祖代效应的评估可能存在低估。本研究结果提醒学界关注环境效应的性别选择性遗传的重要性，并为跨世代选择性传递的近因与远因提出了新的研究问题。

方法
所有行为与形态学数据均以纸质形式现场采集，后转录为电子格式。皮质醇数据通过光密度（Optical Density, OD）值计算得到。