Institute of Tissue Medicine and Pathology, University of Bern, Bern, Switzerland.
Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
Pharmacology. 2024;109(4):216-230. doi: 10.1159/000537864. Epub 2024 Apr 3.
Acute myeloid leukemia (AML) is a cancer of the hematopoietic system characterized by hyperproliferation of undifferentiated cells of the myeloid lineage. While most of AML therapies are focused toward tumor debulking, all-trans retinoic acid (ATRA) induces neutrophil differentiation in the AML subtype acute promyelocytic leukemia (APL). Macroautophagy has been extensively investigated in the context of various cancers and is often dysregulated in AML where it can have context-dependent pro- or anti-leukemogenic effects. On the contrary, the implications of chaperone-mediated autophagy (CMA) on the pathophysiology of diseases are still being explored and its role in AML remains elusive.
We took advantage of human AML primary samples and databases to analyze CMA gene expression and activity. Furthermore, we used ATRA-sensitive (NB4) and -resistant (NB4-R1) APL cells to further dissect a potential function for CMA in ATRA-mediated neutrophil differentiation. NB4-R1 cells are unique in that they do respond to retinoic acid transcriptionally but do not mature in response to retinoid signaling alone unless maturation is triggered by adding cyclic adenosine monophosphate.
Here, we report that CMA-related mRNA transcripts are significantly higher expressed in immature hematopoietic cells as compared to neutrophils, contrasting the macroautophagy gene expression patterns. Accordingly, lysosomal degradation of an mCherry-KFERQ CMA reporter decreases during ATRA-induced differentiation of APL cells. On the other hand, using NB4-R1 cells we found that macroautophagy flux primed ATRA-resistant NB4-R1 cells to differentiate upon ATRA treatment but reduced the association of lysosome-associated membrane protein type 2A (LAMP-2A) and heat shock protein family A (Hsp70) member 8 (HSPA8), necessary for complete neutrophil maturation. Accordingly, depletion of HSPA8 attenuated CMA activity and facilitated APL cell differentiation. In contrast, maintaining high CMA activity by ectopic expression of LAMP-2A impeded APL differentiation.
Overall, our findings suggest that APL neutrophil differentiation requires CMA inactivation and that this pathway predominantly depends on HSPA8 and is possibly assisted by other co-chaperones.
急性髓系白血病（AML）是一种造血系统的癌症，其特征是骨髓谱系未分化细胞的过度增殖。虽然大多数 AML 治疗方法都集中在肿瘤减瘤上，但全反式维甲酸（ATRA）可诱导急性早幼粒细胞白血病（APL）亚型的中性粒细胞分化。自噬在各种癌症的背景下已经被广泛研究，并且在 AML 中经常失调，它可以具有依赖于上下文的促白血病或抗白血病效应。相反，伴侣介导的自噬（CMA）对疾病病理生理学的影响仍在探索中，其在 AML 中的作用仍不清楚。
我们利用人 AML 原代样本和数据库来分析 CMA 基因表达和活性。此外，我们使用 ATRA 敏感（NB4）和 -耐药（NB4-R1）APL 细胞进一步剖析 CMA 在 ATRA 介导的中性粒细胞分化中的潜在功能。NB4-R1 细胞的独特之处在于，它们虽然在转录上对维甲酸有反应，但如果不通过添加环腺苷单磷酸触发成熟，则不会单独对维甲酸信号做出成熟反应。
在这里，我们报告说，与中性粒细胞相比，CMA 相关的 mRNA 转录本在未成熟的造血细胞中表达显著升高，与巨自噬基因表达模式形成对比。相应地，在 APL 细胞的 ATRA 诱导分化过程中，mCherry-KFERQ CMA 报告物的溶酶体降解减少。另一方面，使用 NB4-R1 细胞，我们发现巨自噬通量使 ATRA 耐药的 NB4-R1 细胞在 ATRA 处理后能够分化，但减少了溶酶体相关膜蛋白 2A（LAMP-2A）和热休克蛋白家族 A（Hsp70）成员 8（HSPA8）的结合，这对于完全中性粒细胞成熟是必要的。因此，HSPA8 的耗竭减少了 CMA 活性并促进了 APL 细胞分化。相反，通过过表达 LAMP-2A 维持高 CMA 活性会阻碍 APL 分化。
总体而言，我们的研究结果表明，APL 中性粒细胞分化需要 CMA 失活，该途径主要依赖于 HSPA8，并且可能由其他伴侣协助。
