Abstract
The Awd (abnormal wing discs) gene is the Drosophila homolog of human NME1 and NME2 metastasis suppressor genes. These genes play a key role in tumor progression. Extensive studies revealed that intracellular NME1/2 protein levels could be related to either favorable or poor prognosis depending on tissue context. More recently, extracellular activities of NME1/2 proteins have also been reported, including a tumor- promoting function. We used Drosophila as a genetic model to investigate the mechanism controlling intra- and extracellular levels of NME1/2. We examined the role of several components of the ESCRT (endosomal sorting complex required for transport) complex in controlling Awd trafficking. We show that the Vps28 component of the ESCRT−I complex is required for maintenance of normal intracellular level of Awd in larval adipocytes. We already showed that blocking of Shibire (Shi)/Dynamin function strongly- lowers Awd intracellular level. To further investigate this down regulative effect, we analyzed the distribution of endosomal markers in wild type and Shi-defective adipocytes. Our results suggest that Awd does not enter CD63-positive endosomes. Interestingly, we found that in fat body cells, Awd partly- colocalizes with the ESCRT accessory component ALiX, the ALG-2 (apoptosis-linked gene 2)-interacting protein X. Moreover, we show that the intracellular levels of both proteins are downregulated by blocking the function of the Dynamin encoded by the shibire gene.
Highlights
NME1 and NME2 genes are closely related members of the NME gene family which consists of 10 members (Stafford et al, 2008)
Our studies showed that Awd is an endocytic mediator that interacts with Rab5 and the Drosophila homolog of Dynamin1 encoded by the shibire gene (Dammai et al, 2003; Nallamothu et al, 2008; Woolworth et al, 2009; Ignesti et al, 2014)
The Awd protein is expressed over all the larval fat body that, in 3rd instar larval stage, consists of layers of tightly connected polygonal adipocytes (Figure 1A)
Summary
NME1 and NME2 genes are closely related members of the NME gene family which consists of 10 members (Stafford et al, 2008). Our analyses of the amount and distribution of intracellular Awd in mutant adipocytes highlight a role of Vps28 component of ESCRT–I complex in controlling the Awd presence inside the cell. We found that Awd colocalizes with the ALiX accessory component and that Shi function is required for normal intracellular amount of both proteins.
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