Figure 3

Gene expression analyses for loci at or near the TgPWS/TgAS deletion breakpoints. (A) Tissue expression of Siglec-H. RT-PCR using a cDNA mouse panel shows that Siglec-H (143-bp band) is strongly expressed in adult brain and spleen, with moderate expression in lung and skeletal muscle tissues, but is low or absent in heart, liver, kidney and testis. RNApolII (167-bp) gene expression was used as a control for both (A) and (B). (B) Developmental Siglec-H expression. Very low or absent Siglec-H expression is detected by RT-PCR at E7-E15 with weak expression at late gestational stages (E17). After birth, SiglecH expression is robustly detected in brain at P1 and P4. (C-F) Relative quantification of brain mRNA levels for (C) Siglec-H , (D) Luzp2 , (E) Cezanne2 and (F) Chrna7 was performed by QRT-PCR in two groups of 5 TgPWS (black bars) or 5 TgAS (gray bars) and 5 WT (white bars) mice at P1 and in 4 TgPWS or 4 TgAS and 4 WT mice at P4. All values are presented as the means ± SE: * P ≤ 0.05; ** P ≤ 0.001; *** P ≤ 0.0001, significant differences between WT and TgPWS or TgAS at the indicated time points (Independent samples t-test). (C) Siglec-H mRNA levels are decreased by ~2-fold in TgPWS and TgAS compared with WT at P1 and P4. SiglecH mRNA levels in TgPWS vs. WT were 0.52 ± 0.04 vs. 1.29 ± 0.15 (P = 0.002) and 0.56 ± 0.12 vs. 1.31 ± 0.19 (P = 0.01) at P1 and P4, respectively. Siglec-H mRNA levels were 0.45 ± 0.03 in TgAS compared with 1.18 ± 0.08 in WT at P1 (P = 0.0002) and 0.39 ± 0.04 compared with 0.92 ± 0.07 at P4 (P = 0.0008). (D) Luzp2 mRNA levels in TgPWS vs. WT were 0.98 ± 0.02 vs. 1.25 ± 0.07 (P = 0.009) and 0.91 ± 0.02 vs. 1.11 ± 0.05 (P = 0.01) at P1 and P4, respectively. In TgAS vs. WT mice, mRNA levels were 0.97 ± 0.07 vs. 1.06 ± 0.05 (P = 0.3) at P1 and 0.81 ± 0.05 vs. 0.98 ± 0.02 (P = 0.01) at P4. (E) Cezanne2 shows no difference in expression in TgPWS or TgAS and WT mice. At P1, mRNA levels were 1.01 ± 0.03 for TgPWS vs. 1.04 ± 0.1 for WT (P = 0.8) and 1.03 ± 0.09 for TgAS vs. 1.06 ± 0.03 for WT (P = 0.7). At P4, Cezanne2 mRNA levels were 0.92 ± 0.08 for TgPWS vs. 1.04 ± 0.03 for WT (P = 0.2) and 0.97 ± 0.23 for TgAS vs. 1.09 ± 0.06 for WT (P = 0.6). (F) Chrna7 mRNA levels are ~0.6-fold in TgPWS and TgAS compared with WT mouse brain. At P1 the mRNA levels were 0.64 ± 0.04 in TgPWS vs. 0.99 ± 0.08 in WT (P = 0.005) and 0.57 ± 0.02 in TgAS vs. 0.89 ± 0.1 in WT (P = 0.01), while at P4 mRNA levels for Chrna7 were 0.58 ± 0.01 for TgPWS vs. 1.05 ± 0.03 for WT (P < 0.0001) and 0.63 ± 0.04 for TgAS vs. 1.08 ± 0.12 for WT (P = 0.005).