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N-terminal acetylation and replicative age affect proteasome localization and cell fitness during aging.

van Deventer S, Menendez-Benito V, van Leeuwen F, Neefjes J - J. Cell. Sci. (2014)

Bottom Line: By using a microscopy-based genome-wide screen, we identified genetic factors involved in these processes.Both relocalization of the proteasome and PSG formation were affected by two of the three N-acetylation complexes.These N-acetylation complexes also had different effects on the longevity of cells, indicating that each N-acetylation complex has different roles in proteasome location and aging.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell Biology, Netherlands Cancer Institute and Netherlands Proteomics Center, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.

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In starvation, loss of NatA and NatB has a general effect on reproductive capacity, whereas loss of NatC specifically affects old cells. The reproductive capacity of the different strains in log phase (A) and starvation (B) was assessed by a plating assay. Loss of NatA and NatB compromised reproductive capacity in starved cells, whereas loss of NatC did not. A representative plating assay from three independent experiments is shown. (C) After CFW staining of a starved culture, the cells with the lowest (negative) and highest (positive) CFW signal were sorted to obtain populations of virgin cells and old mothers, respectively. The CFW image is the maximum projection of a 5-µm Z-stack, the DIC image is a single scan in the middle of the Z-stacks. (D) CFUs were counted for old mother (old) and virgin (young) cells. Loss of NatC affects the reproductive capacity of old cells, whereas loss of NatA and NatB reduce the reproductive capacity of both young and old cells. Results are mean±s.d. based on on three independent experiments. Significance was calculated with a paired two-tailed Student's t-test. (* = P<0.05, ** = P<0.01).
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f05: In starvation, loss of NatA and NatB has a general effect on reproductive capacity, whereas loss of NatC specifically affects old cells. The reproductive capacity of the different strains in log phase (A) and starvation (B) was assessed by a plating assay. Loss of NatA and NatB compromised reproductive capacity in starved cells, whereas loss of NatC did not. A representative plating assay from three independent experiments is shown. (C) After CFW staining of a starved culture, the cells with the lowest (negative) and highest (positive) CFW signal were sorted to obtain populations of virgin cells and old mothers, respectively. The CFW image is the maximum projection of a 5-µm Z-stack, the DIC image is a single scan in the middle of the Z-stacks. (D) CFUs were counted for old mother (old) and virgin (young) cells. Loss of NatC affects the reproductive capacity of old cells, whereas loss of NatA and NatB reduce the reproductive capacity of both young and old cells. Results are mean±s.d. based on on three independent experiments. Significance was calculated with a paired two-tailed Student's t-test. (* = P<0.05, ** = P<0.01).

Mentions: As proteasome composition and activity was found to influence longevity in starvation, we wondered whether proteasome localization would also correlate with cell fitness. Cellular fitness in starvation can be determined by assaying the ability of cells to restart their cell cycle when nutrients are added. This is determined by plating equal numbers of cells and quantifying the number of colony-forming units (CFUs). When grown in the presence of sufficient nutrients, the reproductive capacity of NatA- and NatC-deficient cells is similar to WT cells, whereas NatB-deficient cells show a lower reproductive capacity (Fig. 5A) (Polevoda et al., 1999). After a 5-day starvation period, both NatA- and NatB-deficient cells showed lower CFUs than wild-type cells, whereas the reproductive capacity in NatC-deficient cells seemed to be unaffected (Fig. 5B). Proteasome localization in NatA-deficient cells was similar to WT, unlike that in NatB-deficient cells. Given that NatA and NatB deficiency both decrease the number of CFUs measured, proteasome localization cannot be directly related to reproductive capacity in starvation.


N-terminal acetylation and replicative age affect proteasome localization and cell fitness during aging.

van Deventer S, Menendez-Benito V, van Leeuwen F, Neefjes J - J. Cell. Sci. (2014)

In starvation, loss of NatA and NatB has a general effect on reproductive capacity, whereas loss of NatC specifically affects old cells. The reproductive capacity of the different strains in log phase (A) and starvation (B) was assessed by a plating assay. Loss of NatA and NatB compromised reproductive capacity in starved cells, whereas loss of NatC did not. A representative plating assay from three independent experiments is shown. (C) After CFW staining of a starved culture, the cells with the lowest (negative) and highest (positive) CFW signal were sorted to obtain populations of virgin cells and old mothers, respectively. The CFW image is the maximum projection of a 5-µm Z-stack, the DIC image is a single scan in the middle of the Z-stacks. (D) CFUs were counted for old mother (old) and virgin (young) cells. Loss of NatC affects the reproductive capacity of old cells, whereas loss of NatA and NatB reduce the reproductive capacity of both young and old cells. Results are mean±s.d. based on on three independent experiments. Significance was calculated with a paired two-tailed Student's t-test. (* = P<0.05, ** = P<0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4282048&req=5

f05: In starvation, loss of NatA and NatB has a general effect on reproductive capacity, whereas loss of NatC specifically affects old cells. The reproductive capacity of the different strains in log phase (A) and starvation (B) was assessed by a plating assay. Loss of NatA and NatB compromised reproductive capacity in starved cells, whereas loss of NatC did not. A representative plating assay from three independent experiments is shown. (C) After CFW staining of a starved culture, the cells with the lowest (negative) and highest (positive) CFW signal were sorted to obtain populations of virgin cells and old mothers, respectively. The CFW image is the maximum projection of a 5-µm Z-stack, the DIC image is a single scan in the middle of the Z-stacks. (D) CFUs were counted for old mother (old) and virgin (young) cells. Loss of NatC affects the reproductive capacity of old cells, whereas loss of NatA and NatB reduce the reproductive capacity of both young and old cells. Results are mean±s.d. based on on three independent experiments. Significance was calculated with a paired two-tailed Student's t-test. (* = P<0.05, ** = P<0.01).
Mentions: As proteasome composition and activity was found to influence longevity in starvation, we wondered whether proteasome localization would also correlate with cell fitness. Cellular fitness in starvation can be determined by assaying the ability of cells to restart their cell cycle when nutrients are added. This is determined by plating equal numbers of cells and quantifying the number of colony-forming units (CFUs). When grown in the presence of sufficient nutrients, the reproductive capacity of NatA- and NatC-deficient cells is similar to WT cells, whereas NatB-deficient cells show a lower reproductive capacity (Fig. 5A) (Polevoda et al., 1999). After a 5-day starvation period, both NatA- and NatB-deficient cells showed lower CFUs than wild-type cells, whereas the reproductive capacity in NatC-deficient cells seemed to be unaffected (Fig. 5B). Proteasome localization in NatA-deficient cells was similar to WT, unlike that in NatB-deficient cells. Given that NatA and NatB deficiency both decrease the number of CFUs measured, proteasome localization cannot be directly related to reproductive capacity in starvation.

Bottom Line: By using a microscopy-based genome-wide screen, we identified genetic factors involved in these processes.Both relocalization of the proteasome and PSG formation were affected by two of the three N-acetylation complexes.These N-acetylation complexes also had different effects on the longevity of cells, indicating that each N-acetylation complex has different roles in proteasome location and aging.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell Biology, Netherlands Cancer Institute and Netherlands Proteomics Center, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.

Show MeSH
Related in: MedlinePlus