Exploring the Role of Semen Amyloids in Promoting HIV Infection and Fertilization

描述(申请人提供)：HIV最常通过性接触传播，而精液是推动这种致命病毒在全球传播的媒介。我们的研究表明，精液非但不是艾滋病毒的被动载体，反而在体外大大增强了艾滋病毒的感染，我们进一步鉴定了人类精液中的淀粉样纤维，它可以促进艾滋病毒-1与其细胞靶点的融合。这些纤维可以使艾滋病毒感染增加几个数量级以上，因此是开发局部艾滋病毒杀菌剂的良好靶点。这项建议的前两个目标集中在更好地描述这些精液纤维增强艾滋病毒感染性的机制及其对生殖器粘膜内存在的细胞的影响。在目标1中，我建议使用神经生物学领域的技术来表征最有效地增强艾滋病毒感染性的精液纤维的形态特征。这些信息将揭示精液中淀粉样蛋白构象的类型，在努力设计针对精液中宿主因素的特定抑制剂时，应针对这些类型。在目标2中，我将确定精液纤维是否会在宿主细胞中引发炎症，以及这在促进艾滋病毒感染方面可能起到什么作用。众所周知，炎症通常通过招募易感靶细胞和促进HIV基因转录来促进HIV的传播。了解精液纤维在传播过程中对宿主炎症的贡献程度，对于开发有效防止病毒性传播的杀菌剂至关重要。最后，提案的目标3侧重于更好地了解精液原纤维的基本生理功能。精液纤维的进化并不是为了促进艾滋病毒感染，而是可能在人类身上具有生物学目的。有趣的是，HIV与其细胞靶点的融合与精子与卵子的融合有许多相似之处，这增加了这些纤维在受精过程中发挥作用的可能性。为此，我建议在体外受精(IVF)和活体人工授精实验中，确定我们所描述的精液纤维是否促进小鼠配子的融合。了解这些纤维是否有助于促进受精，对于开发艾滋病毒杀微生物剂是至关重要的信息，也可能在生殖领域产生重大影响。已投入大量努力开发一种有效的艾滋病毒杀微生物剂。然而，该领域仍然缺乏一种在预防艾滋病毒性传播方面高度有效的药物，部分原因是我们对围绕粘膜艾滋病毒传播的分子事件缺乏基本了解。这项建议的重点是更好地了解艾滋病毒传播的一个方面，即自然产生的精液纤维增强艾滋病毒传染性的效果。虽然该提议仅限于对这些纤维的体外分析，但我们正在启动在恒河猴身上的平行实验，以检验这些纤维在体内的影响。在K99/R00的指导阶段，如果获得资金，我的计划是开发淀粉样蛋白和生殖生物学的新技术技能，并将这些技能应用于了解艾滋病毒的传播。这将通过与我的K99导师和咨询委员会密切合作来实现。我的长期目标是促进我们对围绕生殖器粘膜中艾滋病毒传播的分子事件的理解，并将这种知识转化为开发新的艾滋病毒预防药物。

DESCRIPTION (provided by applicant): HIV is most frequently transmitted following sexual contact, and semen is the vehicle fueling the global spread of this deadly virus. Far from being a passive vector for HIV, our research revealed that semen drastically enhances HIV infection in vitro, and we have further identified and characterized amyloid fibrils from human semen that increase HIV-1 fusion to its cellular targets. These fibrils can enhance HIV infection by over several orders of magnitude, and therefore serve as good targets for the development of a topical HIV microbicide. The first two aims of this proposal focus on better characterizing the mechanisms by which these semen fibrils enhance HIV infectivity and their influence on cells present within the genital mucosa. In Aim 1, I propose to use techniques from the field of neurobiology to characterize the morphological characteristics of semen fibrils that most effectively enhance HIV infectivity. This information will reveal the types of amyloid conformations in semen that should be targeted in efforts to design specific inhibitors against host factors in semen. In Aim 2, I will determine whether semen fibrils induce inflammation in host cells, and what role this may have in promoting HIV infection. It is known that inflammation generally facilitates HIV transmission by recruiting susceptible target cells and promoting HIV gene transcription. Understanding the extent to which semen fibrils contribute to host inflammation during transmission will be vital for developing microbicides that are effective in preventing sexual transmission of the virus. Lastly, Aim 3 of the proposal focuses on better understanding the fundamental physiological function of semen fibrils. Semen fibrils did not evolve to promote HIV infection, and may have a biological purpose in humans. Intriguingly, HIV fusion to its cellular target shares many properties with the fusion of a spermatozoon to an egg, raising the possibility for a role for these fibrils in fertilization. In this aim, I propose in viro fertilization (IVF) and in vivo artificial insemination experiments to determine if the semen fibris we have characterized promote the fusion of murine gametes. Understanding whether these fibrils serve to promote fertilization is vital information for the development of an HIV microbicide, and could also have a significant impact in the field of reproduction. Substantial efforts have been invested into developing an effective HIV microbicide. However, the field still lacks a drug that is highly effective at preventing the sexual spread of HIV, in part due to our lack of basic understanding of the molecular events surrounding mucosal HIV transmission. This proposal focuses on better understanding one aspect of HIV transmission, namely the effect of naturally-occurring semen fibrils that enhance HIV infectivity. Although the proposal is limited to in vitro analysis of these fibrils, we are initiating experiments in parallel in rhesus macaques to examine the effect of these fibrils in vivo. My plan during the mentored phase of the K99/R00, were it to get funded, is to develop new technical skills in amyloid and reproductive biology and to apply these skills towards understanding HIV transmission. This will be accomplished by working closely with my K99 mentors and advisory committee. My long-term goal is to advance our understanding of the molecular events surrounding HIV transmission in the genital mucosa, and to translate this knowledge into the development of new classes of HIV preventatives.