Prostate cancer : Low level of syntaphilin (SNPH) increase risk of metastasis

Modulation of mitochondrial dynamics by SNPH ubiquitination. A, One frame of 82 from a confocal microscopy image sequence showing mitotracker-labeled mitochondria (top). Mitochondria are segmented in each frame and tracked throughout the image sequence (bottom). The colored mesh indicates the segmentation boundaries, with different colors and numeric labels assigned to different tracked mitochondria. Tracking results are used to identify fission and fusion events, with track initiation indicating mitochondrial fusion and track termination indicating mitochondrial fission events. B, Mitochondrial dynamics was assessed by time-lapse videomicroscopy in Yumm 1.7 cells transfected with vector, WT SNPH or SNPH-mutant K111R or K153R. Representative images of 3D rendering of individual cells per condition are shown. C, The conditions are as in A and B, and mitochondrial fusion and fission events/voxel/frame were calculated in Yumm 1.7 transfectants. Data are the mean ± SEM (n = 61 movies from 10 imaging experiments after deconvolution: Vector, 9 movies; WT SNPH, 20 movies; SNPH-mutant K111R, 19 movies; SNPH-mutant K154R, 13 movies). D, PC3 cells were transfected with WT SNPH or SNPH-mutant K111R or K153R and total cell extracts (TCE) or mitochondrial (Mito) fractions were analyzed by Western blotting. E, PC3 cells were transfected with control nontargeting siRNA (Ctrl) or SNPH-directed siRNA and cytosol (Cyto) or mitochondrial (Mito) extracts were analyzed by Western blotting. F, PC3 cells were transfected with nontargeting siRNA (Ctrl) or CHIP- or USP7-directed siRNA and total cell extracts were analyzed by Western blotting. For D and F, antibodies to the different Drp1 phosphorylation (p) sites are indicated.

For about 90 percent of men with prostate cancer, the cancer remains localized to the primary site, resulting in a five-year survival rate of almost 100 percent.

Unfortunately, the remaining 10 percent of patients develop locally invasive and metastatic disease, which increases the severity of the disease and likelihood of death and limits treatment options.

A report in The American Journal of Pathology indicates that a significantly lower presence of syntaphilin (SNPH) – a mitochondrial protein – within the tumor’s central core versus at the tumor’s invasive outer edge, may identify patients at increased risk of metastasis.

Although most tumors reprogram their metabolism towards glycolysis even when oxygen is present, that is the “Warburg effect” (1), there is now mounting evidence that mitochondria continue to play a fundamental role in cancer (2, 3). This is especially important in advanced disease, where mitochondrial function has been linked to tumor repopulation after oncogene ablation (4), drug resistance (5), and tumor progression in vivo (6). In addition, mitochondria have been implicated in tumor cell movements and metastasis (7) through modulation of oxidative metabolism (8), organelle biogenesis (8), buffering of oxidative stress (9), and cycles of organelle fusion and fission, that is dynamics (10).

In this context, recent evidence has demonstrated that in response to stress stimuli of the microenvironment, tumor cells reposition mitochondria from their constitutive perinuclear localization to the peripheral (or cortical) cytoskeleton (11).

In turn, these cortical mitochondria function as a “spatiotemporal” energy source to fuel membrane lamellipodia dynamics, kinase signaling and actin cytoskeleton remodeling, promoting increased cellular chemotaxis and invasion (11–13).

There are intriguing similarities between these observations in cancer (11–13) and the process of mitochondrial trafficking in neurons, which also repositions mitochondria at sites of high energy demands (14).

These suggest that tumors may hijack mitochondrial trafficking as a bioenergetics requirement of cell motility, especially under stress (11), facilitating metastatic dissemination in vivo.

Accordingly, a genome-wide siRNA screen has recently identified syntaphilin (SNPH), a molecule known to inhibit mitochondrial trafficking in neurons (15), as a regulator of tumor cell movements (16).

Although considered neuronal-specific, SNPH is, in fact, broadly expressed in cancer, suppresses mitochondrial trafficking to the cortical cytoskeleton, and restricts tumor cell movements and metastasis in vivo (16).

In addition, analysis of genomic databases suggests that SNPH is progressively downregulated or lost during tumor progression, correlating with worse disease outcome (16).

How this pathway is regulated, however, has remained elusive. In this study, we examined a role of SNPH ubiquitination in the control of mitochondrial trafficking and metastasis.

Syntaphilin (SNPH) inhibits the movement of mitochondria in tumor cells, preventing their accumulation at the cortical cytoskeleton and limiting the bioenergetics of cell motility and invasion.

Although this may suppress metastasis, the regulation of the SNPH pathway is not well understood.

Using a global proteomics screen, we show that SNPH associates with multiple regulators of ubiquitin-dependent responses and is ubiquitinated by the E3 ligase CHIP (or STUB1) on Lys111 and Lys153 in the microtubule-binding domain.

SNPH ubiquitination did not result in protein degradation, but instead anchored SNPH on tubulin to inhibit mitochondrial motility and cycles of organelle fusion and fission, that is dynamics.

Expression of ubiquitination-defective SNPH mutant Lys111→Arg or Lys153→Arg increased the speed and distance traveled by mitochondria, repositioned mitochondria to the cortical cytoskeleton, and supported heightened tumor chemotaxis, invasion, and metastasis in vivo.

Interference with SNPH ubiquitination activated mitochondrial dynamics, resulting in increased recruitment of the fission regulator dynamin-related protein-1 (Drp1) to mitochondria and Drp1-dependent tumor cell motility.

These data uncover nondegradative ubiquitination of SNPH as a key regulator of mitochondrial trafficking and tumor cell motility and invasion.

In this way, SNPH may function as a unique, ubiquitination-regulated suppressor of metastasis.

These patients may require more rigorous testing, surveillance, and treatment.

“Predicting aggressive behavior in prostate cancer is an entirely unmet and urgent need.

There are currently no tissue-based biomarkers to help clinicians reliably identify the subset of prostate cancer patients who will progress to life-threatening, disseminated disease and who would, therefore, benefit from systemic therapies before or following prostatectomy.

If our findings are supported by larger studies, SNPH measurement in tumors could be developed into a predictive biomarker,” explained Marie E. Robert, MD, of the Department of Pathology, Yale School of Medicine, New Haven, CT, USA.

(A) Low magnification view of cancer nodule with the central tumor bulk consisting predominantly of invasive cribriform Gleason pattern 4 (hematoxylin and eosin staining). (B) Low magnification view of the same cancer nodule with an outer rim of accentuated syntaphilin (SNPH) staining. The invasive front is defined as the external rim of each cancer nodule abutting the adjacent stroma as outlined in this image as the zone between the dotted and solid lines. Arrows denote accentuated SNPH staining at the tumor-stromal interface. Credit: The American Journal of Pathology

In this study, investigators found that SNPH, a key determinant of the balance between tumor cell proliferation and tumor cell invasion, is abundantly expressed in prostate cancer, where it exhibits high expression at the invasive tumor edge compared to the central tumor bulk, correlating with greater cell proliferation at the tumor edge location.

They also found that SNPH expression increases with increasing Gleason pattern.

Of potential vital clinical relevance, low central tumor expression correlated with an increased risk of metastasis in a group of patients undergoing radical prostatectomy.

The investigators analyzed tissue specimens from 89 men with prostate adenocarcinoma who had undergone removal of the prostate.

They found that SNPH manifests a unique, biphasic spatial distribution in prostate tumors, meaning that in 96 percent of the prostate cancer tumors analyzed, SNPH levels were elevated at the outer invasive edge where it correlates with increased tumor cell proliferation, but decreased within the central tumor core.

This differential was more pronounced in more advanced tumors.

Importantly, central tumor SNPH measures (H scores) were significantly lower in 16 patients with metastases compared to patients without metastases, whereas SNPH scores at the invasive edges were not different in patients with or without metastases.

Most of the metastases also expressed SNPH strongly.

The researchers suggest that SNPH acts as a negative regulator of mitochondrial activity and that its down-regulation in the central portions of primary prostate cancer is associated with an increased risk of metastatic disease.

“This is the first study to suggest a clinical role for SNPH assessment in prostate cancer prognosis, potentially confirming recent evidence in experimental models of its importance in the phenotypic switch between proliferative and metastatic tumor states.

Our results also reaffirm a critical, emerging role of mitochondrial biology in influencing tumor behavior,” Dr. Robert noted.

More information: “Syntaphilin Is a Novel, Biphasic Biomarker of Aggressive Prostate Cancer and a Metastasis Predictor,” American Journal of Pathology (2019). DOI: 10.1016/j.ajpath.2019.02.009
Journal information: American Journal of Pathology
Provided by Elsevier


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