Tau mRNA and protein expression in E10+/− and wild-type mice. (A) Only 3R-tau mRNA (212 bp) was expressed at birth (P1) in wild-type mice (wt). At (P9), 3R-tau and 4R-tau mRNA (305 bp) was expressed, while at (P18) and in 2-month-old wt-mice, exon 10 was included and 4R tau mRNA was expressed. In contrast, 2-month-old E10+/− mice (E10) produced 3R-tau and 4R-tau mRNA in equal proportions. Signals depended on mRNA and reverse transcriptase in the cDNA synthesis reaction (−RT, E10). (B) Left; Western blot with a selective antibody for 3R tau resulted in a 38 kDa band in 2 months-old E10+/− mouse brain (E10). Right; Two additional bands were seen when films were overexposed. These bands likely represented tau-isoforms with N-terminal domains exon 2 and 3. In an age-matched wt-mouse there was only a small amount of 3R-tau protein. (C) Left; An antibody (Tau-1) that recognizes 3R-tau as well as 4R-tau generated two protein bands at 38 and 39 kDa in brains of 2-month-old E10+/− mice (E10). Right; Overexposure results in two additional double bands likely representing isoforms of tau with the N-terminal domains exon 2 and 3 of tau protein. Only a small amount of 3R tau protein was found in age-matched wild-type littermates (wt). Recombinant human tau protein representing all tau isoforms and a newborn wild-type mouse brain (P1) was included as a positive control. (D) Immunostaining with selective antibody 3RD showed that 3R-tau was mainly localized in axons of E10+/− mice, (E) but not present in an age-matched wild-type mice. (F-G) The non-selective antibody Tau-1, which recognizes both 3R-tau and 4R-tau, stained hippocampal layers which are rich in axons in both E10+/− (F) and wild-type mice (G). (H-I) No immunostaining in E10+/− (H) or wild-type mice in the absence of primary antibody (I).
Gumucio et al. BMC Neuroscience 2013 14:148 doi:10.1186/1471-2202-14-148