Fig. 1

Effect of loss of Camk4 on the turnover and PTMs of Tf as well as iron content in different mouse tissues. a: Immunohistochemical staining of Tf promoter-trapped GFP expression in mouse brain. Data obtained from GENSAT database. B-D: RNA in situ hybridization images showing an abundance of Tfrc (b), Camkk2 (c), and Camk4 (d) mRNAs in mouse brain. Images obtained from BGEM database [37]. Red arrows indicate overlapping gene expressions in the cerebellum region. e: Immunoblots showing expression of Camkk2 and Gapdh in the cerebellum and cortex tissues form adult wild-type and Camkk2^−/− mice. f-g: Immunoblots showing expression of Camk4 and Erk1/2 in the cerebellum and liver tissues from adult wild-type and Camk4^−/− mice. Ns: nonspecific. h-i: Immunoblots showing expression of Tf and Tfrc in the cerebellum and liver tissues from wild-type and Camk4^−/− mice. Bottom panels represent oriole-stained gel showing total protein loading. Relative intensities of the red arrow-marked oriole-stained bands in the total protein profile were used for the quantification of Tf and Tfrc. j-m: Scatter plots showing the relative abundance of Tf and Tfrc in the cerebellum and liver tissues. N = 2 replicates from three wild-type and Camk4^−/− mice. P values by t-test (unpaired). n: Scatter plot showing iron content in different mouse tissues. N = 26/27, ~ 9 replicates from three wild-type and Camk4^−/− mice respectively. P values by one-way ANOVA followed by Dunnett’s multiple comparison test. o & r: Immunoblots showing charged fractions of Tf, separated by IEF/SDS-PAGE. Colored dotted rectangles indicate alterations in the differentially charged fractions of Tf. (P&S): Overlapped plot profiles of the immunoblots presented in o & r respetively. In the cerebellum, p100-245 kDa Tf spots were considered for plot profiling. In the liver tissue, p75-100 kDa Tf spots were considered for plot profiling. q &t: Scatter plots showing the relative amount of negatively charged fraction of Tf (red rectangle areas). N = 6, 2 replicates from three wild-type and Camk4^−/− mice respectively. P values by t-test (unpaired).