DNA Damage Triggers Golgi Dispersal via GOLPH3 and DNA-PK
- Author(s): Farber-Katz, Suzette Elena
- Advisor(s): Field, Seth
- et al.
Golgi morphology and function has fascinated scientists for decades; however, it was unclear how the two were related. Our research delving into the function of GOLPH3 provides evidence for how Golgi morphology relates to Golgi trafficking. We identified GOLPH3 as a novel PtdIns(4)P-binding protein. We found that GOLPH3 requires PtdIns(4)P for its localization at the Golgi. We also showed that GOLPH3 binds to MYO18A, which results in a tensile force that pulls vesicles off the trans-Golgi. We showed that GOLPH3, MYO18A, and F-actin are all required for trafficking from the Golgi. The tensile force also stretches the Golgi and creates the Golgi ribbon that extends around the nucleus. Therefore, we identified a novel mechanism that explains how Golgi trafficking results in the extended Golgi ribbon.
While we have an understanding of how GOLPH3 functions, it was unclear how GOLPH3 is regulated. To better understand its regulation, we performed immunoprecipitation-mass spectrometry experiments to identify its phosphorylation sites. We found that T143 and T148 were phosphorylated. By analyzing the sequences surrounding these sites, we noticed that T143 and Q144 comprise a "TQ" motif, which are recognized by kinases in the DNA damage pathway. Thus, we examined whether DNA damage has any effect on Golgi morphology, and we found that it causes Golgi dispersal. We showed that DNA damage-induced Golgi dispersal requires phosphorylation of GOLPH3, as well as MYO18A and F-actin. Our extensive data argue that DNA-PK is the kinase that phosphorylates GOLPH3 on the TQ motif. We also demonstrated that DNA damage increases the interaction between GOLPH3 and MYO18A, thereby increasing the tensile force pulling on the Golgi membranes to elicit a dispersed Golgi. Because GOLPH3 was previously identified as an oncogene, we measured survival after DNA damage, and we found that GOLPH3, MYO18A, and DNA-PK are required for survival after DNA damage. Furthermore, knockdown of GOLPH3 or MYO18A increases apoptosis levels after DNA damage. Our data provide new insight into how the DNA-PK/GOLPH3/MYO18A/F-actin pathway causes dramatic Golgi reorganization. These results will have implications for cancer therapy, as inhibition of this pathway in the presence of DNA damage strikingly increases cell death.