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The Regulatory Factor ZFHX3 Modifies Circadian Function in SCN via an AT Motif-Driven Axis.

Parsons MJ, Brancaccio M, Sethi S, Maywood ES, Satija R, Edwards JK, Jagannath A, Couch Y, Finelli MJ, Smyllie NJ, Esapa C, Butler R, Barnard AR, Chesham JE, Saito S, Joynson G, Wells S, Foster RG, Oliver PL, Simon MM, Mallon AM, Hastings MH, Nolan PM - Cell (2015)

Bottom Line: Using RNA sequencing, we found minimal effects on core clock genes in Zfhx3(Sci/+) SCN, whereas the expression of neuropeptides critical for SCN intercellular signaling was significantly disturbed.Lentiviral transduction of SCN slices showed that the ZFHX3-mediated activation of AT motifs is circadian, with decreased amplitude and robustness of these oscillations in Zfhx3(Sci/+) SCN slices.In conclusion, by cloning Zfhx3(Sci), we have uncovered a circadian transcriptional axis that determines the period and robustness of behavioral and SCN molecular rhythms.

View Article: PubMed Central - PubMed

Affiliation: MRC Harwell, Harwell Science and Innovation Campus, Oxfordshire OX11 0RD, UK.

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AVP and VIP Immunofluorescence Co-localizes with ZFHX3 Immunofluorescence, Related to Figure 4(A and B) Confocal micrographs showing co-immunofluorescence of ZFHX3 and (A) AVP or (B) VIP within the SCN (scale bar, 100 μm) and at high magnification (scale bar, 20 μm) at ZT4.(C) Both AVP (blue bar) and VIP (red bar) neurons almost completely co-localized with ZFHX3 (n = 3 brains), graph shows mean ± SEM for the percentage of co-localized cells.
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figs3: AVP and VIP Immunofluorescence Co-localizes with ZFHX3 Immunofluorescence, Related to Figure 4(A and B) Confocal micrographs showing co-immunofluorescence of ZFHX3 and (A) AVP or (B) VIP within the SCN (scale bar, 100 μm) and at high magnification (scale bar, 20 μm) at ZT4.(C) Both AVP (blue bar) and VIP (red bar) neurons almost completely co-localized with ZFHX3 (n = 3 brains), graph shows mean ± SEM for the percentage of co-localized cells.

Mentions: Using immunofluorescence, we compared AVP, GRP, VIP, and VIPR2 levels in Zfhx3Sci/+ and Zfhx3+/+ SCN. We found that GRP and VIP levels, but not AVP and VIPR2 levels, were significantly decreased in Zfhx3Sci/+ at ZT6 (Figures 4E–4I). There was a small but significant decrease in Zfhx3Sci/+ SCN size compared to Zfhx3+/+ (Zfhx3Sci/+ SCN size: 53,600 ± 2,400 μm2, n = 4; Zfhx3+/+ SCN size: 66,900 ± 1,600 μm2, n = 4; p < 0.05, t test, area of DAPI signal fluorescence). The data suggest that ZFHX3 actively influences the expression of these clock-relevant neuropeptides in the adult SCN and that this activation is disrupted in Zfhx3Sci/+ mice. These differences were not primarily due to aberrant terminal differentiation, as there was no significant reduction in the number of AVP or VIP immuno-positive neurons (Figures 4J and 4K). Interestingly, ZFHX3 immuno-positive neurons almost completely overlapped with those for both AVP and VIP (Figure S3).


The Regulatory Factor ZFHX3 Modifies Circadian Function in SCN via an AT Motif-Driven Axis.

Parsons MJ, Brancaccio M, Sethi S, Maywood ES, Satija R, Edwards JK, Jagannath A, Couch Y, Finelli MJ, Smyllie NJ, Esapa C, Butler R, Barnard AR, Chesham JE, Saito S, Joynson G, Wells S, Foster RG, Oliver PL, Simon MM, Mallon AM, Hastings MH, Nolan PM - Cell (2015)

AVP and VIP Immunofluorescence Co-localizes with ZFHX3 Immunofluorescence, Related to Figure 4(A and B) Confocal micrographs showing co-immunofluorescence of ZFHX3 and (A) AVP or (B) VIP within the SCN (scale bar, 100 μm) and at high magnification (scale bar, 20 μm) at ZT4.(C) Both AVP (blue bar) and VIP (red bar) neurons almost completely co-localized with ZFHX3 (n = 3 brains), graph shows mean ± SEM for the percentage of co-localized cells.
© Copyright Policy - CC BY
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4537516&req=5

figs3: AVP and VIP Immunofluorescence Co-localizes with ZFHX3 Immunofluorescence, Related to Figure 4(A and B) Confocal micrographs showing co-immunofluorescence of ZFHX3 and (A) AVP or (B) VIP within the SCN (scale bar, 100 μm) and at high magnification (scale bar, 20 μm) at ZT4.(C) Both AVP (blue bar) and VIP (red bar) neurons almost completely co-localized with ZFHX3 (n = 3 brains), graph shows mean ± SEM for the percentage of co-localized cells.
Mentions: Using immunofluorescence, we compared AVP, GRP, VIP, and VIPR2 levels in Zfhx3Sci/+ and Zfhx3+/+ SCN. We found that GRP and VIP levels, but not AVP and VIPR2 levels, were significantly decreased in Zfhx3Sci/+ at ZT6 (Figures 4E–4I). There was a small but significant decrease in Zfhx3Sci/+ SCN size compared to Zfhx3+/+ (Zfhx3Sci/+ SCN size: 53,600 ± 2,400 μm2, n = 4; Zfhx3+/+ SCN size: 66,900 ± 1,600 μm2, n = 4; p < 0.05, t test, area of DAPI signal fluorescence). The data suggest that ZFHX3 actively influences the expression of these clock-relevant neuropeptides in the adult SCN and that this activation is disrupted in Zfhx3Sci/+ mice. These differences were not primarily due to aberrant terminal differentiation, as there was no significant reduction in the number of AVP or VIP immuno-positive neurons (Figures 4J and 4K). Interestingly, ZFHX3 immuno-positive neurons almost completely overlapped with those for both AVP and VIP (Figure S3).

Bottom Line: Using RNA sequencing, we found minimal effects on core clock genes in Zfhx3(Sci/+) SCN, whereas the expression of neuropeptides critical for SCN intercellular signaling was significantly disturbed.Lentiviral transduction of SCN slices showed that the ZFHX3-mediated activation of AT motifs is circadian, with decreased amplitude and robustness of these oscillations in Zfhx3(Sci/+) SCN slices.In conclusion, by cloning Zfhx3(Sci), we have uncovered a circadian transcriptional axis that determines the period and robustness of behavioral and SCN molecular rhythms.

View Article: PubMed Central - PubMed

Affiliation: MRC Harwell, Harwell Science and Innovation Campus, Oxfordshire OX11 0RD, UK.

Show MeSH
Related in: MedlinePlus