Recent studies have shown that loss of pollen-S function in S₄' pollen from sweet cherry (Prunus avium) is associated with a mutation in an S haplotype-specific F-box4 (SFB4) gene. However, how this mutation leads to self-compatibility is unclear. Here, we examined this mechanism by analyzing several self-compatible sweet cherry varieties. We determined that mutated SFB4 (SFB4') in S4' pollen (pollen harboring the SFB4' gene) is approximately 6 kD shorter than wild-type SFB4 due to a premature termination caused by a four-nucleotide deletion. SFB4' did not interact with S-RNase. However, a protein in S4' pollen ubiquitinated S-RNase, resulting in its degradation via the 26S proteasome pathway, indicating that factors in S4' pollen other than SFB4 participate in S-RNase recognition and degradation. To identify these factors, we used S₄-RNase as a bait to screen S4' pollen proteins. Our screen identified the protein encoded by S₄-SLFL2, a low-polymorphic gene that is closely linked to the S-locus. Further investigations indicate that SLFL2 ubiquitinates S-RNase, leading to its degradation. Subcellular localization analysis showed that SFB4 is primarily localized to the pollen tube tip, whereas SLFL2 is not. When S₄-SLFL2 expression was suppressed by antisense oligonucleotide treatment in wild-type pollen tubes, pollen still had the capacity to ubiquitinate S-RNase; however, this ubiquitin-labeled S-RNase was not degraded via the 26S proteasome pathway, suggesting that SFB4 does not participate in the degradation of S-RNase. When SFB4 loses its function, S₄-SLFL2 might mediate the ubiquitination and degradation of S-RNase, which is consistent with the self-compatibility of S4' pollen.

最近的研究表明，S中花粉-S功能的丧失₄甜樱桃'花粉（甜樱桃) 与 S 单倍型特异性 F-box4 (SFB4) 基因的突变有关。然而，这种突变如何导致自我相容性尚不清楚。在这里，我们通过分析几个自交的甜樱桃品种来研究这种机制。我们确定 S4' 花粉（含有 SFB4' 基因的花粉）中的突变 SFB4 (SFB4') 比野生型 SFB4 短约 6 kD，这是由于四核苷酸缺失引起的过早终止。SFB4' 不与 S-RNase 相互作用。然而，S4' 花粉中的一种蛋白质泛素化 S-RNase，导致其通过 26S 蛋白酶体途径降解，表明 S4' 花粉中除 SFB4 之外的其他因子参与了 S-RNase 的识别和降解。为了识别这些因素，我们使用了 S ₄-RNase 作为诱饵筛选 S4' 花粉蛋白。我们的筛选鉴定了由S ₄ -SLFL2编码的蛋白质，这是一种与 S 基因座密切相关的低多态性基因。进一步的研究表明 SLFL2 泛素化 S-RNase，导致其降解。亚细胞定位分析表明，SFB4 主要定位于花粉管尖端，而 SLFL2 则不是。当S ₄ -SLFL2表达在野生型花粉管中被反义寡核苷酸处理抑制时，花粉仍然具有泛素化S-RNase 的能力；然而，这种泛素标记的 S-RNase 并未通过 26S 蛋白酶体途径降解，这表明 SFB4 不参与 S-RNase 的降解。当SFB4失去作用时，S₄ -SLFL2 可能介导 S-RNase 的泛素化和降解，这与 S4' 花粉的自交亲和性一致。