Understand the stages of mitophagy. Definition of subcellular localization. Discover the articles on these subjects recently published in the journal Molecular and cellular proteomics.
Work from outside to
As cells adapt to the stresses of life, mitochondria must also adapt. Over time, mitochondria are damaged and then recycled through a specialized autophagy process called mitophagy.
Zittlau et al.
Shown over time from left to right, mitochondria are broken down from the outside
During mitophagy, proteins are marked for degradation by ubiquitination. Phosphorylation works in tandem with ubiquitination, as each modification can provide either positive or negative feedback to the other. In particular, mitophagy requires parkin E3 ubiquitin ligase and serine/threonine-protein kinase PINK1. Researchers have identified mutations in both enzymes in Parkinson’s disease, so understanding their function is critical.
Despite an abundant literature, scientists still have key questions about the process of mitophagy. Some researchers have demonstrated that autophagosomes consume mitochondria at the same time during mitophagy, although others suggest that individual sections within mitochondria can be broken down piece by piece.
In a recent paper published in the journal Molecular and Cellular Proteomics, Katharina I. Zittlau and colleagues from the University of Tübingen describe the use of a three-level proteomics approach to examine parkin-dependent mitophagy in HeLa cells. After inducing mitophagy, the team quantified total protein levels over 18 hours to assess mitochondrial protein degradation in the presence or absence of functional parkin. They also measured changes in protein ubiquitination and phosphorylation.
Their data support an outer-inner breakdown of mitochondria during mitophagy, showing evidence of protein ubiquitination and degradation in outer mitochondrial compartments first, with inner compartments following later. Using a large dataset, the researchers also identified instances in which a phosphorylation event blocked or enhanced ubiquitination during mitophagy. In particular, they showed that dephosphorylation of voltage-gated anion-selective channel protein 2 is required for its parkin-dependent ubiquitination and ultimate degradation.
This study provides detailed information that improves our knowledge of mitophagy as well as the specific contribution of parkin at each step of the process.
The cytoplasm is a dense but well-organized space. A multitude of biochemical reactions occur simultaneously, each localized in a specific domain, such as inside the nucleus, on the mitochondrial surface or at the level of the plasma membrane. Accordingly, the localization of RNA and proteins – as well as intracellular trafficking to their final destination – must be tightly regulated. Disrupting this spatial organization can be problematic, as many diseases are characterized by poorly localized proteins.
Although researchers can gain a wealth of knowledge from global transcriptomic and proteomic datasets, the story is incomplete without spatial information. Local enrichment or depletion of macromolecules—often masked in whole-cell omics—regulates these biological pathways and, ultimately, cellular function.
Josie A. Christopher and a team from the University of Cambridge recently published a comprehensive study exam methods used to study the subcellular localization of proteins and RNA in the review Molecular and cellular proteomics. The authors provide a detailed overview of techniques such as microscopy-based assays, imaging mass cytometry, and proteomic/transcriptomic coupling with biochemical fractionations or proximity labeling. The advantages and limitations of each are discussed to help readers select the best methods for their own projects.
The techniques highlighted in this review will be crucial in answering fundamental questions about cellular organization and paving the way for translational research and new diagnostic approaches.