Anton Simeonov Ph.D. National Institute of Health

Asymmetric Division of Old Mitochondria During Replication Maintains Stem Cell Phenotype

As cells grow and mature, they accumulate defective building blocks in the form of oxidized, unfolded, or otherwise covalently damaged proteins, as well as partially defective organelles. The work by Katajisto et al. provides novel ways of monitoring the fate of defective organelles as the cells divide into two daughter cells. Of particular interest to the field has been the purported segregation of aged mitochondria between the two daughter cells in dividing stem cells. The team fused photoactivatable green fluorescent protein (paGFP) to various targeting signals or proteins in order to monitor lysosomes, mitochondria, the Golgi apparatus, ribosomes, and chromatin. Not only does paGFP fluoresce exquisitely after exposure to a UV light pulse, but since its biosynthesis continues after the light pulse, one has the ability to follow the fate of unlabeled “young” components as well the labeled “old” components, which can appear segregated or co-localized within individual cells. As a model, the authors used stem-like cells (SLCs) present in immortalized human mammary epithelial cells, which carry a distinct round shape morphology making them easy to distinguish from the flat nonstem-like mammary epithelial counterparts (figure above). The team found that unlike regular epithelial cells in which all analyzed cellular components segregated symmetrically, the SLCs displayed bias in apportioning old mitochondria with respect to each daughter cell. Using the Snap-tag technology, which allows in-cell covalent labeling of specific proteins with fluorophores of choice, the fate of the old and young mitochondria could be followed (figure below). It was shown that old mitochondria were consistently segregated across generations, thus leading to their relative dilution in the new stem-like progeny, with the progeny containing more of the old mitochondria being more likely to lose its stem-like properties and to differentiate. The asymmetric apportionment was associated with preferential perinuclear localization of old mitochondria, in contrast to the young organelles, which appeared to be distributed more evenly. Thus, it appears that a quality control mechanism exists which forces old mitochondria carrying a lower membrane potential to assume perinuclear localization, which in turn allows the young mitochondria carrying a higher membrane potential (known to be important for maintenance of stemness) to be enriched spatially during division. In closing, the authors surmise that a similar mechanism may have a bearing on a related phenomena of cell aging in old organisms, making it important to study further ways to control this process of asymmetric mitochondria apportioning.

Age-dependent segregation and subcellular localization of mitochondria. (A) Schematic of the labeling strategy using Snap-tag chemistry. (B) and (C) Analysis of mitochondrial outer membrane (B) and inner membrane (C) inheritance upon cell division. Red and green sections of bars represent the old and young labels, respectively. Values were scaled so that total intensity (red + green) of the mother cell is 1 (n = 5). Representative division occurring at 6 hours after the second (green) label is shown. Percent values represent the average of five divisions. Original magnification 40×. (D) Confocal microscopy of a cell with ≥50-hour-old and 0- to 1-hour-old mitochondrial Snap-Omp25 labeled red and green, respectively. Mitochondrial network contains domains with different levels of enrichment for the old proteins. Mitochondrial domains enriched with old proteins (arrowheads) are not associated with autophagosomes detected by immunofluorescence for LC3B (purple) (63×, 2 μm Z section). (E) Localization of old (red) and young (green) mitochondria (Snap-Omp25) 10 hours after labeling in an undivided cell. Squares mark regions used for measurements of the perinuclear and peripheral intensities in frames captured 10 hours after labeling for n = 3 (cells imaged from three separate labeling experiments) (*P < 0.05, **P < 0.01, t test).

* Abstract from Science 2015; Vol. 348, p. 340–343

By dividing asymmetrically, stem cells can generate two daughter cells with distinct fates. However, evidence is limited in mammalian systems for the selective apportioning of subcellular contents between daughters. We followed the fates of old and young organelles during the division of human mammary stemlike cells and found that such cells apportion aged mitochondria asymmetrically between daughter cells. Daughter cells that received fewer old mitochondria maintained stem cell traits. Inhibition of mitochondrial fission disrupted both the age-dependent subcellular localization and segregation of mitochondria and caused loss of stem cell properties in the progeny cells. Hence, mechanisms exist for mammalian stemlike cells to asymmetrically sort aged and young mitochondria, and these are important for maintaining stemness properties.

Anton Simeonov, Ph.D., works at the NIH

ASSAY & Drug Development Technologies, published by Mary Ann Liebert, Inc., offers a unique combination of original research and reports on the techniques and tools being used in cutting-edge drug development. The journal includes a "Literature Search and Review" column that identifies published papers of note and discusses their importance. GEN presents here one article that was analyzed in the "Literature Search and Review" column, a paper published in Science titled "Asymmetric apportioning of aged mitochondria between daughter cells is required for stemness." Authors of the paper are Katajisto P, Döhla J, Chaffer CL, Pentinmikko N, Marjanovic N, Iqbal S, Zoncu R, Chen W, Weinberg RA, Sabatini DM