parg1-1 is a long circadian period mutant
The lack of effect of PARP and SRT mutants on circadian period might be unexpected because the tej-1 mutation ofPARG1 , which is predicted to encode a poly(ADPribose) glycohydrolase that counter acts PARP activity has been reported to increase the period of circadian rhythms in Arabidopsis (Panda et al ., 2002). We therefore reinvestigated whether lesions in PARG1can affect circadian period. As reported previously, tej-1 (a point mutation which results in a change of the glycine at position 262 to a glutamic acid) increases circadian period (Supplemental Figure 5a, C24 26.8 ± 0.6 h, tej -1 30.5 ± 2.64 h, P = 0.006). Having confirmed the phenotype of the tej-1 mutant, we next sought to confirm whether the phenotype of tej-1 was due to a lesion inPARG1. We obtained a second allele of PARG1(At2g31870), we named as parg1-1 (SALK_147805, NASC; Figure 5a). When transformed with the CCA1::LUC reporter,parg1 -1 mutants had a slightly longer circadian period of luminescence (Supplemental Figure 5b, Col-0 24.5 ± 0.6 h, parg1 -1 24.8 ± 0.4 h, P = 0.021). The mean period in circadian rhythms of leaf movement was 1 - 1.5 hours longer than wild type (Figure 5b, Col-0 24.3 ± 1.0 h, parg1-1 25.4 ± 1.3 h, P < 0.001). Similarly, circadian rhythms in DF were long period in the parg1-1 mutant (Figure 5c, Col-0 23.4 ± 0.9 h, parg1-1 25.0 ± 1.1 h, P = 0.011). We also isolated insertion mutants of PARG2 (At2g31865),parg2-1 (GK072_B04, NASC), to determine if this close gene family member might also contribute to circadian regulation (Figure 5a). The parg2-1 mutant by contrast had weak to no effect on circadian rhythms of leaf movement (Figure 5d, Col-0 24.5 ± 0.7 h, parg2-124.8 ± 0.6 h, P = 0.209) or delayed chlorophyll fluorescence (Figure 5e, Col-0 24.23 ± 3.06 h, parg2 -1 23.2 ± 0.8 h, P = 0.667). Thus, the effects of the PARG1 mutant are specific to that member of the gene family.