Critical Review 1

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Discovering an old DoGs’ new trick

Heterotrimeric G proteins regulate a variety of signaling pathways that control cell development and influ- ence cell morphology via actin/cyto- skeleton remodeling. There are four main families of G proteins: Gi/Go, Gq, Gs and G12/13. Researchers long have thought that Gs, unlike its family members, is coupled specifically and exclusively to adenylyl cyclases.

In a new study published in the Journal of Biological Chemistry, Alejandro Castillo–Kauil of the Center for Research and Advanced Studies of the National Polytechnic Institute and collaborators challenge this dogmatic view by identifying a new Gs target. Using biochemical, molecular biological and chemo- genetic approaches, the researchers demonstrated that the Gαs subfamily of G proteins can regulate the activity of Rho GTPases such as Rho guanine nucleotide exchange factor, or Rho- GEF. The interaction identified by the group activates the small G protein Cdc42 by Gs-coupled GPCRs, stimu- lating a rearrangement of the cyto- skeleton and inducing formation of fingerlike protrusions called filopodia.

These results provide new insight into G protein activity and define a new role for RhoGEF coupling in G protein function. DOI: 10.1074/jbc.AC120.015204

A pathogen’s proteins target mitochondria

The tick-borne pathogen Coxiella burnetii causes Q fever, or query fever, a rare flulike disease that can spread to humans who inhale dust particles contaminated by infected farm or

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Noninvasive tool provides oral cancer prognosis Oral squamous cell carcinoma, which affects about 34,000 people

in the U.S. each year, is found in the cells lining the lips and mouth. Metastasis to the lymph nodes is a sign of disease progression and may be accompanied by changes in proteolytic activity. During proteolysis, enzymes cut up proteins into short fragments called peptides. Recent work suggests that characterizing the sequence and abundance of these molecules — a method dubbed peptidomics — might provide new in- sight on cancer biology and in the clinic. In a recent paper in the journal Molecular & Cellular Proteomics, Leandro Xavier Neves of the Brazil- ian Biosciences National Laboratory and a team of Brazilian clinicians and scientists describe their analysis of oral squamous cell carcinoma patient saliva using peptidomics.

After extracting peptides from saliva samples, the research team ana- lyzed and compared the peptide content in samples from patients with and without metastasis to the lymph nodes. They found more than 1,000 uniquely expressed peptides in each group and an additional 1,628 pep- tides expressed by both groups. A series of statistical analyses identified 77 peptides of particular interest; all of these peptides are overexpressed in samples from patients with lymph node metastasis, which supports the hypothesis that proteolytic activity increases in metastatic disease. Ten of these peptides also correlated with clinical features of metastatic tumors. The researchers used a panel measuring the abundance of five peptides to classify patients according to metastatic state.

To predict which enzymes generated this distinctive saliva peptidome, the team compared the sequences of peptides and full-length proteins to identify the most common cleavage sites. Many of the predicted en- zymes are associated with the lysosome or vacuole, linked to immunity or present in the oral microbiome. They then examined the expression

of these enzymes in publicly available oral squamous cell carcinoma tissue data. This analysis showed that enhanced expression of enzymes — but not the proteins they target — associates with metastasis and reduced survival, further bolster- ing the link between proteolytic activity and disease progression.

Together, these analy- ses of proteolysis in oral

squamous cell carcinoma may be useful for patient prognosis. DOI: 10.1074/mcp.RA120.002227 — Nuala Del Piccolo

CDC/UNSPLASH

Peptidomics analyses of saliva samples from oral squamous cell carcinoma patients correlate with disease metastasis. The technique may be a useful noninvasive prognostic tool.

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FEBRUARY 2021 ASBMB TODAY 27

domestic animals. When C. burnetii infects a human macrophage, it transfers as many as 150 effector proteins into the host cell. Research- ers know these proteins change the host’s physiology to support the infection. However, they know little about the contributions of the indi- vidual effector proteins, partly due to their diversity and scarcity.

Laura Fielden and colleagues at the University of Melbourne hy- pothesized that C. burnetii effector proteins target the mitochondria — known as the powerhouse of the cell — because that organelle is crucial in cellular homeostasis. To test this, the team isolated mitochondria from human macrophages infected with C. burnetii and analyzed the proteins using high-sensitivity mass spectrometry.

The team’s recent paper in the journal Molecular & Cellular Proteomics identified seven C. burnetii proteins associated with the mitochondria, including two not previously found in the organ- elle. They found that one of these effector proteins, MceC, moves to the inner mitochondrial membrane and interacts with mitochondrial proteins involved in quality control and metabolic function.

This study presents an adapt- able method for characterizing the contributions of a pathogen’s effector proteins. DOI: 10.1074/mcp.RA120.002370

Pinpointing therapies in PAK pathways

The serine/threonine kinase P21-activated kinase-1, or PAK1, influences cancer-related biologi- cal processes such as cell migration, invasion and angiogenesis. However, researchers have not completely delineated the network of signaling

molecules linked to PAK1. Jae-Hong Kim of Kyungpook

National University and a team of in- ternational collaborators used genetic transformation screens, RNAi, phar- macological inhibition and migration assays to characterize PAK1-mediated signal transduction pathways thor- oughly. The researchers selected 19 candidate PAK1 genetic interactions that had human orthologs and were expressed in glioma for further ex- amination in mammalian cells, brain slice cultures and orthotopic glioma models. RNAi and pharmacological inhibition of potential PAK1 interac- tors confirmed the importance of several genes related to the mitotic spindle, proteolysis, autophagy and metabolism in PAK1-mediated glioma cell migration, drug resistance and proliferation.

These results, published in the Journal of Biological Chemistry, provide a comprehensive view of PAK1-mediated signal transduction pathways and identify new drug targets for glioma therapy. DOI: 10.1074/jbc.RA120.014831

Nuala Del Piccolo (ndelpiccolo@ ucdavis.edu) is a science writer in the biomedical engineering department at the University of California, Davis. She earned her Ph.D. in materials science and engineering at Johns Hopkins University.

Anand Rao ([email protected]) is an ASBMB science communi- cator. Follow him on Twitter @AnandRaoPhD.

Lisa Nicole Learman (llearman@ jhmi.edu) is a Ph.D. candidate studying molecular neurosci- ence at Johns Hopkins University School of Medicine. Follow her on Twitter @LearmanLisa.

Laurel Oldach (loldach@ asbmb.org) is a science writer for the ASBMB. Follow her on Twitter @LaurelOld.

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