Distinct Biochemical Pools of Golgi Phosphoprotein 3 in the Human Breast Cancer Cell Lines MCF7 and MDA-MB-231

Abstract

Golgi phosphoprotein 3 (GOLPH3) has been implicated in the development of carcinomas in many human tissues, and is currently considered a bona fide oncoprotein. Importantly, several tumor types show overexpression of GOLPH3, which is associated with tumor progress and poor prognosis. However, the underlying molecular mechanisms that connect GOLPH3 function with tumorigenicity are poorly understood. Experimental evidence shows that depletion of GOLPH3 abolishes transformation and proliferation of tumor cells in GOLPH3-overexpressing cell lines. Conversely, GOLPH3 overexpression drives transformation of primary cell lines and enhances mouse xenograft tumor growth in vivo. This evidence suggests that overexpression of GOLPH3 could result in distinct features of GOLPH3 in tumor cells compared to that of non-tumorigenic cells. GOLPH3 is a peripheral membrane protein mostly localized at the trans-Golgi network, and its association with Golgi membranes depends on binding to phosphatidylinositol-4-phosphate. GOLPH3 is also contained in a large cytosolic pool that rapidly exchanges with Golgi-associated pools. GOLPH3 has also been observed associated with vesicles and tubules arising from the Golgi, as well as other cellular compartments, and hence it has been implicated in several membrane trafficking events. Whether these and other features are typical to all different types of cells is unknown. Moreover, it remains undetermined how GOLPH3 acts as an oncoprotein at the Golgi. Therefore, to better understand the roles of GOLPH3 in cancer cells, we sought to compare some of its biochemical and cellular properties in the human breast cancer cell lines MCF7 and MDA-MB-231 with that of the non-tumorigenic breast human cell line MCF 10A. We found unexpected differences that support the notion that in different cancer cells, overexpression of GOLPH3 functions in diverse fashions, which may influence specific tumorigenic phenotypes.

FAP-a and GOLPH3 Are Hallmarks of DCIS Progression to Invasive Breast Cancer

“…Kenneth L. Scott raised the hypothesis that GOLPH3 interacted with VPS35 and the tetramer to recycle receptor for key molecules, thus regulating downstream mTOR signaling (19). Tenorio et al found that the cytosolic and membrane components of the three breast cell lines had biochemical differences in GOLPH3, that was that, in cancer cells, part of the overexpressed GOLPH3 was modified by differentiation (34).…”

Autophagosomes cooperate in the degradation of intracellular C‐terminal fragments of the amyloid precursor protein via the MVB/lysosomal pathway

“…After the final wash, coverslips were mounted onto glass slides with Fluoromount‐G (SouthernBiotech, Birmingham, AL, USA). Images of fixed cells were acquired with an AxioObserver.D1 microscope equipped with a PlanApo ×63 oil‐immersion objective (NA 1.4) and an AxioCam MRm digital camera (Carl Zeiss GmbH, Jena, Germany), using similar settings described previously (34), or with an Olympus FluoView FV1000 scanning unit fitted on an inverted Olympus IX81 microscope equipped with a PlanApo ×60 oil‐immersion objective (NA 1.4; Olympus, Tokyo, Japan), using similar settings as described previously (35). Alternatively, images of fixed cells were acquired by a DeltaVision OMX system (GE Healthcare Life Sciences, Issaquah, WA, USA) for superresolution by structured illumination microscopy (SIM), using similar settings as described previously (36, 37).…”

“…The corrected fluorescent signal in each cell of each image was used to determine the total integrated pixel intensity per cell area by ImageJ 1.44o software [Image Processing and Analysis in Java; National Institutes of Health (NIH), Bethesda, MD, USA; http://imagej.nih.gov/]. Colocalization analyses and estimation of the Pearson's correlation coefficient (38) were performed as described in Wong and Holzbauer (35).…”

Oncogenic Roles of GOLPH3 in the Physiopathology of Cancer

“…To investigate how the overexpression of GOLPH3 might influence its roles in the oncogenesis of breast cancer, the group of Mardones compared some of the biochemical and cellular characteristics of GOLPH3 in the two human breast cancer cell lines MCF7 and MDA-MB-231 with those of GOLPH3 in the non-tumorigenic human breast cell line MCF 10A [81]. They showed that the three breast cell lines possess biochemical distinct pools of GOLPH3 and that the GOLPH3 protein distributes differently in cytosolic and membrane-bound pools in the different breast cell lines.…”