Fig 2: Radiation-induced long-term changes in IGF1, IGFBP3, and IGF1R. A) Serum IGF1 levels at 2 months post-exposure, B) Serum IGFBP3 levels at 2 months post-exposure, C) Molar ratio of IGF1/IGFBP3 representing free IGF1 levels at 2 months post-exposure, D) Serum IGF1 levels at 12 months post-exposure, E) Serum IGFBP3 levels at 12 months post-exposure, F) Molar ratio of IFG1/IGFBP3 at 12 months post-exposure. G) Representative images of IGF1R immunostained small intestine sections at 12 months post-irradiation. H) Quantitation of IGF1R signal intensity in small intestine sections. I) Representative images of IGF1R immunostained colon sections at 12 months post-irradiation. J) Quantitation of IGF1R signal intensity in colon sections. Error bars represent mean ± SEM and p<0.05 was considered significant, compared to sham-irradiated control.

Fig 3: TMEM263 deficiency results in marked reduction in circulating insulin-like growth factor 1 (IGF-1), IGF binding protein 3 (IGFBP3), and IGF acid labile subunit (IGFALS) levels.Serum levels of growth hormone (GH; A), IGF-1 (B), IGFBP3 (C), IGFALS (D), insulin (E), glucose (F), calcium (G), and phosphate (H) in wild-type (WT) (+/+), heterozygous (+/-), and Tmem263-KO (-/-) male and female mice at 8 weeks of age. Sample size for panel A (GH): males (WT = 9; het = 9; KO = 5) and females (WT = 10; het = 7; KO = 8). Panel B (IGF-1): males (WT = 9; het = 9; KO = 9) and females (WT = 10; het = 7; KO = 7). Panel C (IGFBP3): males (WT = 15; het = 16; KO = 11) and females (WT = 12; het = 18; KO = 10). Panel D (IGFALS): males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). Panel E (insulin): males (WT = 12; het = 17; KO = 8) and females (WT = 10; het = 16; KO = 8). Panel F (glucose): males (WT = 9; het = 14; KO = 9) and females (WT = 6; het = 15; KO = 8). Panel G and H (calcium and phosphate): males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). (I) Ratio of calcium-to-phosphate in WT, heterozygous, and KO male and female mice. Sample size for males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). All data are presented as mean ± SEM. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 (one-way ANOVA with Tukey’s multiple comparisons test).

Fig 4: Reduced hepatic growth hormone receptor (GHR) protein level and signaling in TMEM263 knockout (KO) mice.(A) The growth hormone (GH)/insulin-like growth factor 1 (IGF-1) axis required for postnatal skeletal growth. At the onset of growth spurt, growth hormone releasing hormone (GHRH) from the hypothalamus causes the release of GH from the anterior pituitary. Circulating GH binds to its receptor (GHR) in liver and other peripheral tissues to induce the synthesis and secretion of IGF-1, which then acts in an endocrine, paracrine, and/or autocrine manner to induce skeletal growth. (B) Expression levels of Ghr (growth hormone receptor), Igf1, Igfals (IGF binding protein acid labile subunit), and Igfbp3 (IGF binding protein 3) transcripts in the liver of wild-type (WT) and KO mice. Sample size of male mice (WT, n=8; KO, n=8) and female mice (WT, n=8; KO, n=8). (C) Immunoblot analysis of GHR protein levels in the liver of WT (n=7) and KO (n=7) mice. Molecular weight markers are indicated on the left. (D) Quantification of the immunoblot results as shown in C (n=7 per genotype). (E–F) Reduced hepatic GH-induced signaling in KO (-/-; n=5) mice relative to WT (+/+; n=4) controls. Immunoblot analysis of phospho-JAK2 (Tyr1008), total JAK2, phospho-STAT5 (Y694), and total STAT5 in liver lysates from control male mice not injected with GH (E) and male mice injected with recombinant GH (F). Molecular weight markers are indicated on the left of the gel. (G) Quantification of the immunoblot results as shown in F (WT, n=4; KO, n=5). All data are presented as mean ± SEM. **p<0.01; ***p<0.001; ****p<0.0001 (one-way ANOVA with Tukey’s multiple comparisons test for data in B and two-tailed Student’s t-test for data in D and F). Figure 5—source data 1.Top left – Original uncropped membrane from imager showing blue channel only as a black and white image.Two close molecular markers are noted. Growth hormone receptor (GHR) at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-GHR antibody. Top middle – Original uncropped membrane from imager showing blue channel only as black and white. Two molecular markers are noted. β-Actin at the predicted molecular weight is marked. The cropped region used for the main figure is marked with a dotted box. The membrane was probed with the anti-β-actin antibody. Top right – The same membrane as the top middle, but overexposed and imaged with the light channel to show the entire membrane outline. The anti-β-actin antibody is very clean and therefore the membrane boarder is hard to see otherwise. Bottom left – Original uncropped membrane from imager showing blue channel only as a black and white image. Two close molecular markers are noted. GHR at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-GHR antibody. Bottom middle – Original uncropped membrane from imager showing blue channel only as black and white. Two molecular markers are noted. β-Actin at the predicted molecular weight is marked. The cropped region used for the main figure is marked with a dotted box. The membrane was probed with the anti-β-actin antibody. Bottom right – The same membrane as the top middle, but overexposed and imaged with the light channel to show the entire membrane outline. The anti-β-actin antibody is very clean and therefore the membrane boarder is hard to see otherwise. Figure 5—source data 2.Top left – Original uncropped membrane from imager showing blue channel only as a black and white image.Two close molecular markers are noted. Phospho-JAK2 at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-phospho-JAK2 antibody. Top right – The same membrane as on the left, re-probed to visualize total-JAK2 signal. Bottom left – Original uncropped membrane from imager showing blue channel only as a black and white image. Two close molecular markers are noted. Phospho-STAT5b at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-phospho-STAT5b antibody. Bottom right – The same membrane as on the bottom left, re-probed to visualize total-STAT5b signal. Figure 5—source data 3.Top left – Original uncropped membrane from imager showing blue channel only as a black and white image.Two close molecular markers are noted. Phospho-JAK2 at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-phospho-JAK2 antibody. Bottom left – The same membrane as on the top left, re-probed to visualize total-JAK2 signal. Top right – Original uncropped membrane from imager showing blue channel only as a black and white image. Two close molecular markers are noted. Phospho-STAT5b at the predicted molecular weight is marked. The cropped region used for the main figure image is marked with a dotted box. This membrane was probed with the anti-phospho-STAT5 antibody. Bottom right – The same membrane as on the top right, re-probed to visualize total-STAT5b signal.