Figure 4: SIRT6 vacates L1 promoters in aged cells leading to derepression of L1. | Nature Communications

Figure 4: SIRT6 vacates L1 promoters in aged cells leading to derepression of L1.

From: SIRT6 represses LINE1 retrotransposons by ribosylating KAP1 but this repression fails with stress and age

Figure 4

(a) Senescent cells exhibit elevated L1 expression. Total cellular RNA was isolated from young and replicatively senescent HDF cells. L1 levels were quantified by qRT–PCR and normalized to actin expression; n=5, error bars indicate s.d. (b) Brain and liver tissue from old mice exhibits increased L1 expression. Total RNA was isolated from the indicated tissues from young (4 months old) and old (24 months old) mice. L1 levels were quantified by qRT–PCR and normalized to actin expression; n=3, error bars indicate s.d.; NS, not significant. (c) SIRT6 is depleted from the L1 5′-UTR in senescent cells. ChIP with SIRT6 antibodies revealed that the protein is depleted from the L1 5′-UTR in senescent cells, relative to young cells. Relative enrichment, after normalization to input and H3 levels is shown; n=3, error bars indicate s.d. (d) SIRT6 is depleted from the L1 5′-UTR in old brain tissue. ChIP with SIRT6 antibodies revealed that the protein is depleted from the L1 5′-UTR in old brain tissue, relative to young brain tissue. Relative enrichment, after normalization to input and H3 levels is shown; n=3, error bars indicate s.d. (e) Overexpression of SIRT6 partially restores L1 expression to youthful levels in senescent cells. Young and senescent cells were transfected with either a HRPT- or SIRT6-encoding expression plasmid. Total RNA was isolated from these cells and L1 expression was quantified by qRT–PCR after normalization to actin expression; n=3, error bars indicate s.d. (f) DNA damage is sufficient to rapidly induce SIRT6 relocalization. ChIP-qPCR with SIRT6 antibodies revealed that the protein is rapidly depleted from the L1 5′-UTR in response to gamma irradiation. (g) Overexpression of SIRT6 attenuates gamma irradiation induced activation of L1 transcription. Cells were transfected with the indicated vector and then irradiated with gamma irradiation. L1 levels were quantified by qRT–PCR; n=3, error bars indicate s.d.; NS, not significant. Where appropriate, statistical significance was determined by the Student’s t-test.

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