Exploring the impact of night shift work on methylation of circadian genes

Figures & data

Table 1. Demographic data among current day and night shift study participants.

	Current day worker (n = 38)	Current night shift worker (n = 36)
Age (years) (n, %)
<40	7 (18.4)	16 (44.4)
40–54	24 (63.2)	11 (30.6)
≥55	7 (18.4)	9 (25.0)
Menopausal status (n, %)
Postmenopausal	12 (31.6)	14 (38.9)
Premenopausal	26 (68.4)	22 (61.1)
Education (n, %)
High school	2 (5.3)	0 (0.0)
Post-secondary diploma	19 (50.0)	19 (52.8)
University undergraduate degree or higher	17 (44.8)	17 (47.2)
Household Income (in Canadian $) (n, %)
<$75,000	10 (26.3)	3 (8.3)
$75,000-$99,999	9 (23.7)	8 (22.2)
≥$100,000	19 (50.0)	25 (69.4)
Smoking status (n, %)
Never smoked	24 (63.2)	25 (69.4)
Past smoker	10 (26.3)	9 (25.0)
Currently smoke/recently quit	4 (10.5)	2 (5.6)
Family history of cancer (n, %)
Yes	17 (44.7)	19 (52.7)
No	21 (55.3)	16 (44.4)
Missing	0 (0.0)	1 (2.8)
Alcohol consumption (n, %)
≥2+ drinks/week	10 (26.3)	10 (27.7)
1–4 drinks/month	18 (47.4)	15 (41.7)
Never or <1 drink/month	10 (26.3)	11 (30.6)
Occupational exposure to radiation (n, %)
Not often	32 (84.2)	19 (52.8)
Very often/often	6 (15.8)	17 (47.2)
Occupational exposure to disinfectants (n, %)
Not often	15 (39.5)	1 (2.8)
Very often/often	23 (60.5)	35 (97.2)
White blood cell-type composition (mean, SD)
B cells (%)	2.73 (1.40)	3.59 (1.98)
CD8 T cells (%)	6.91 (2.95)	6.95 (3.92)
CD4 T cells (%)	18.47 (4.95)	19.51 (6.40)
Natural killer cells (%)	3.14 (2.24)	2.59 (2.41)
Monocytes (%)	6.92 (2.50)	6.63 (2.16)
Neutrophils (%)	61.31 (7.40)	59.96 (7.56)
Characteristics of work schedule
Status (n, %)
Part-time	5 (13.2)	5 (13.9)
Full-time	33 (86.8)	31 (86.1)
Night shift work duration (n, %)
<10 years	26 (68.4)	10 (27.8)
≥10 years	12 (31.6)	26 (72.2)
Night shift work intensity (n, %)
<3 consecutive night shifts/week	-	25 (69.4)
≥3 consecutive night shifts/week	-	11 (30.6)

Table 2. Circadian gene methylation differences between current night shift and day shift workers.

Illumina ID	Chromosome	Position	Gene	Gene Group^a	Relation to CpG Island^b	Mean^c methylation in day workers (SD)	Mean^c methylation in night shift workers (SD)	Adjusted^d mean difference (95% CI)	q-value^e
cg14718583	22	38,725,060	CSNK1E	5ʹUTR	Open sea	87.72 (1.84)	88.57 (1.57)	1.69 (0.87, 2.52)	0.18
cg20667664	17	38,254,448	NR1D1	Body	North Shore	78.97 (1.65)	78.29 (1.51)	−1.53 (−2.38, −0.67)	0.52
cg12596843	11	45,864,825	CRY2	-	North Shelf	12.73 (1.28)	12.41 (0.84)	−0.89 (−1.43, −0.35)	0.67
cg23929615	22	38,794,688	CSNK1E	TSS200	Island	52.41 (2.28)	51.06 (2.09)	−2.07 (−3.34, −0.79)	0.67
cg10472395	15	61,070,548	RORA	Body	Open Sea	82.50 (1.38)	81.96 (1.45)	−1.09 (−1.80, −0.39)	0.69
cg10976861	2	239,149,937	PER2	TSS1500	South Shore	33.86 (2.11)	32.64 (1.61)	−1.58 (−2.60, −0.55)	0.69
cg26113056	15	61,521,643	RORA	TSS200	Island	8.97 (1.56)	8.57 (1.21)	−1.03 (−1.71, −0.35)	0.69
cg23965982	15	61,055,848	RORA	Body	Open Sea	83.96 (1.91)	83.44 (1.41)	−1.23 (−2.07, −0.38)	0.78
cg16774421	12	27,481,875	ARNTL2	-	North Shelf	92.20 (1.33)	91.85 (1.14)	−0.91 (−1.54, −0.27)	0.78
cg13184823	4	187,476,599	MTNR1A	TSS200	Island	38.96 (8.07)	41.24 (6.26)	5.42 (1.59, 9.25)	0.78

a. Functional region of gene as indicated in Illumina annotation.

b. Position relative to CpG Island as indicated in Illumina annotation.

c. Unadjusted mean.

d. All analyses adjusted for age, income, part/full-time status, occupational exposures to radiation and disinfectants, and leukocyte cell profile. Day workers are the referent category.

e. Q-values represent p-values adjusted for multiple testing using the FDR method.

Table 3. Circadian gene methylation differences between workers with ≥10 years of night shift work duration and workers with <10 years.

Illumina ID	Chromosome	Position	Gene	Gene Group^a	Relation to CpG Island^b	Mean^c methylation in <10 years workers (SD)	Mean^c methylation in ≥10 years workers (SD)	Adjusted^d mean difference (95% CI)	q-value^e
cg20667664	17	38,254,448	NR1D1	Body	North Shore	79.13 (1.73)	78.15 (1.33)	−1.60 (−2.33 – 0.87)	0.08
cg22405816	9	118,135,937	DEC1	5ʹUTR	Open Sea	84.68 (2.47)	83.25 (2.65)	−1.84 (−2.87,0.81)	0.32
cg00436663	22	38,793,933	CSNK1E	Body	Island	7.54 (0.93)	7.07 (0.85)	−0.74 (−1.18, – 0.31)	0.32
cg24219929	15	60,884,748	RORA	Body	Island	9.83 (7.03)	6.37 (3.38)	−4.81 (−7.63, – 1.98)	0.32
cg18204040	12	26,279,190	BHLHE41	TSS1500	Island	10.69 (0.72)	10.20 (0.69)	−0.56 (−0.90, −0.22)	0.32
cg06233947	16	58,230,251	CSNK2A2	Body	North Shore	92.20 (1.17)	91.32 (1.49)	−1.13 (−1.81, – 0.44)	0.32
cg26724232	15	60,885,912	RORA	Body	South Shore	81.46 (3.30)	83.61 (2.65)	2.24 (0.85, 3.63)	0.32
cg09473510	4	187,476,573	MTNR1A	TSS200	Island	19.92 (8.21)	23.99 (7.45)	5.19 (1.97, 8.41)	0.32
cg13154331	22	38,795,868	CSNK1E	TSS1500	South Shore	70.52 (2.44)	71.55 (2.17)	1.52 (0.57, 2.48)	0.32
cg07250429	15	61,051,202	RORA	Body	Open Sea	84.76 (1.22)	83.98 (1.57)	−1.03 (−1.68, –0.38)	0.32

a. Functional region of gene as indicated in Illumina annotation.

b. Position relative to CpG Island as indicated in Illumina annotation.

c. Unadjusted mean.

d. All analyses adjusted for age, income, part/full-time status, occupational exposures to radiation and disinfectants, and leukocyte cell profile. Workers with <10 years of night shift work duration are the referent category.

e. Q-values represent p-values adjusted for multiple testing using the FDR method.

Table 4. Circadian gene methylation differences between workers with ≥3 consecutive night shifts/week and <3 consecutive night shifts/week.

Illumina ID	Chromo-some	Position	Gene	Gene Group^a	Relation to CpG Island^b	Mean^c methylation in <3 shifts/week workers (SD)	Mean^c methylation in ≥3 shifts/week workers (SD)	Adjusted^d mean difference (95% CI)	q-value^e
cg21078679	11	13,377,829	ARNTL	Body	Open sea	85.79 (1.36)	87.71 (2.14)	2.21 (1.15, 3.27)	0.15
cg26449680	22	38,714,272	CSNK1E	TSS200	South Shore	37.84 (2.92)	40.81 (2.69)	1.71 (0.65, 2.77)	0.97
cg15603424	11	13,300,592	ARNTL	5ʹUTR	Island	8.10 (4.32)	12.34 (5.28)	4.72 (1.65, 7.79)	0.97
cg14718583	22	38,725,060	CSNK1E	5ʹUTR	Open Sea	87.97 (1.76)	89.07 (1.48)	1.65 (0.56, 2.75)	0.97
cg27004243	17	8,055,360	PER1	5ʹUTR	Island	6.00 (0.64)	5.45 (0.87)	−0.75 (−1.24, −0.25)	0.97
cg03092603	22	38,712,210	CSNK1E	5ʹUTR	North Shore	7.10 (0.72)	6.52 (0.60)	−0.72 (−1.22, −0.22)	0.97
cg23633210	2	101,600,602	NPAS2	Body	Open Sea	86.66 (1.58)	87.94 (1.36)	1.47 (0.43, 2.51)	0.97
cg17367616	12	27,485,428	ARNTL2	TSS1500	North Shore	56.72 (2.93)	55.81 (3.87)	−2.56 (−4.48, −0.65)	0.97
cg04324336	15	60,941,269	RORA	Body	Open Sea	86.00 (1.45)	84.89 (1.48)	−1.14 (−2.11, −0.18)	0.97
cg08924113	12	27,497,913	ARNTL2	Body	Open Sea	79.10 (4.68)	76.00 (7.06)	−4.30 (−7.94, −0.66)	0.97

a. Functional region of gene as indicated in Illumina annotation.

b. Position relative to CpG Island as indicated in Illumina annotation.

c. Unadjusted mean.

d. All analyses adjusted for age, income, part/full-time status, occupational exposures to radiation and disinfectants, and leukocyte cell profile. Workers with <3 consecutive night shifts/week are the referent category.

e. Q-values represent p-values adjusted for multiple testing using the FDR method.

Figure 1. Volcano plot of results from the analysis of 1150 CpG loci across 22 circadian genes in association with A) current night shift work vs. day work, B) night shift work duration (≥10 years vs. <10 years) and C) night shift intensity (≥3 consecutive night shifts/week vs. <3 night shifts/week). The figure plots p-values versus the effect size (adjusted mean difference in methylation β-values). Dotted line represents p = 0.05. Loci with FDR ≤0.20 are highlighted.