Disabling of ARC1 through CRISPR-Cas9 leads to a complete breakdown of self-incompatibility responses in Brassica napus.

Disabling of ARC1 through CRISPR-Cas9 leads to a complete breakdown of self-incompatibility responses in Brassica napus.
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通过 CRISPR/CAS9 禁用 ARC1 导致甘蓝型油菜自交不亲和反应完全崩溃

DOI:
10.1016/j.xplc.2022.100504
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发表时间:
2023-03-13
影响因子:
10.5
通讯作者:
Samuel MA
Samuel MA
中科院分区:
生物学1区
文献类型:
--
作者:
Abhinandan K;Hickerson NMN;Lan X;Samuel MA

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许多开花植物利用自交不亲和性(SI)反应作为遗传机制来防止自花花粉在柱头上形成,从而促进异交和遗传多样性。在芸苔属 SI 期间,S 位点受体激酶 (SRK) 识别花粉配体 SP11 会导致 E3 连接酶 ARM 重复序列蛋白 (ARC1) 的激活,从而导致成功花粉接受所需的相容性因子被蛋白酶体降解。 ARC1 最初被鉴定为 SRK 激酶结构域的相互作用因子,并在成熟柱头中高度表达 (Gu et al., 1998)。 ARC1 的反义抑制导致自交不亲和甘蓝型油菜 W1 系中 SI 部分分解,确立了 ARC1 作为 SI 正调节因子的作用(Stone 等人,1999)。在自交不亲和的琴叶草中,RNAi 介导的 ARC1 抑制进一步支持了这一点,导致 SI 途径部分崩溃 (Indriolo et al., 2012)。在芸苔属和拟南芥中观察到的 SI 部分受损表明,替代的 SI 途径或 ARC1 的不完全抑制可能导致 SI 的不完全破坏。这个问题仍然悬而未决。在这项研究中,我们创建了 ARC1 功能丧失的甘蓝型油菜品系,以证明 ARC1 在介导 SI 反应中的核心作用。尽管在两个不同的系统中显示 SI 部分崩溃,但 ARC1 在十字花科 SI 反应中的作用在过去几十年里一直受到质疑。芸苔属 SLG、SRK 和 ARC1 的稳定共表达不足以在相容的拟南芥中赋予 SI 表型(Bi 等,2000)。在另一份报告中,当拟南芥植物被转化以表达 SRKb 和 SCRb 基因时,在没有 ARC1 的情况下观察到强烈的 SI 表型(Kitashiba 等人,2011)。仅通过 SRK 和 SCR 转化表现出 SI 表型的拟南芥植物在遗传力、发育调控和 SI 响应强度方面表现出与自交不亲和芸苔属品种相似的趋势(Nasrallah 和 Nasrallah,2014)。尽管在进化上,缺乏真正的 ARC1 直系同源物的拟南芥可以重新利用其他 E3 连接酶来承担 ARC1 的角色,但 ARC1 的完全缺失如何影响芸苔属物种中的 SI 的问题。仍未解决。
Many flowering plants utilize the self-incompatibility (SI) response as a genetic mechanism to prevent self-pollen from establishing on the stigmas, thereby promoting outcrossing and genetic diversity. In Brassica during SI, recognition of the pollen ligand SP11 by S-locus receptor kinase (SRK) results in activation of the E3 ligase ARM-Repeat-Containing protein (ARC1), which leads to proteasomal degradation of compatibility factors required for successful pollen acceptance. ARC1 was originally identified as an interactor of the SRK kinase domain and is highly expressed in mature stigmas (Gu et al., 1998). Antisense suppression of ARC1 resulted in partial breakdown of SI in the selfincompatible Brassica napus W1 line, establishing the role of ARC1 as a positive regulator of SI (Stone et al., 1999). This was further supported by RNAi-mediated suppression of ARC1 in the self-incompatible A. lyrata, which resulted in partial breakdown of the SI pathway (Indriolo et al., 2012). The observed partial compromise of SI in both Brassica and Arabidopsis suggested that either an alternative SI pathway or incomplete suppression of ARC1 could have resulted in the incomplete breakdown of SI. This question has remained unresolved. In this study, we created ARC1 loss-of-function B. napus lines to demonstrate the central role of ARC1 in mediating the SI response.Despite the partial breakdown of SI shown in two different systems, the role of ARC1 in the Brassicaceae SI response has been questioned for the past several decades. Stable coexpression of Brassica SLG, SRK, and ARC1 was insufficient to confer the SI phenotype in compatible Arabidopsis thaliana (Bi et al., 2000). In another report, when A. thaliana plants were transformed to express SRKb and SCRb genes, a strong SI phenotype was observed in the absence of ARC1 (Kitashiba et al., 2011). A. thaliana plants that exhibit the SI phenotype through transformation of SRK and SCR alone show trends in heritability, developmental regulation, and intensity of SI response similar to those of self-incompatible Brassica cultivars (Nasrallah and Nasrallah, 2014). Although, evolutionarily, Arabidopsis that lacked a bona fide ARC1 ortholog could have re-purposed other E3 ligases to assume the role of ARC1, the question of how complete loss of ARC1 could influence SI in Brassica sp. remained unresolved.
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