Effects of Sortilin Inhibition with Alexa Fluor-488 siRNA in TM4 Sertoli Cells

摘要

Sortilin, a 95kDa type I sorting receptor has been implicated in the lysosomal trafficking of sphingolipid activator proteins. Several soluble lysosomal hydrolases, including various cathepsins, reach the lysosomes in absence of the mannose 6-phosphate receptor (M6P-Rc). The M6P-Rc is considered the main lysosomal sorting receptor of mammalian cells. Based on these observations, we postulated that sortilin operates as an alternative receptor for these hydrolases. Thus, our objective was to study the effect of sortilin inhibition on the transport of two cathepsins in the TM4 Sertoli cell line. TM4 cells were transfected with a sortilin siRNA labeled with Alexa Fluor-488 and viewed by confocal microscopy. Our results showed that the expression of sortilin was abolished by the siRNA. The immunostaining with cathepsin D and H showed a granular staining in the non-transfected cells and was significantly decreased in the transfected cells. These results indicate that sortilin inhibition affected the transport of cathepsin D and H to the lysosomes and implicate sortilin in the sorting of soluble hydrolases.