B-raf Alternative Splicing Is Dispensable for Development but Required for Learning and Memory Associated with the Hippocampus in the Adult Mouse

Abstract

The B-raf proto-oncogene exerts essential functions during development and adulthood. It is required for various processes, such as placental development, postnatal nervous system myelination and adult learning and memory. The mouse B-raf gene encodes several isoforms resulting from alternative splicing of exons 8b and 9b located in the hinge region upstream of the kinase domain. These alternative sequences modulate the biochemical and biological properties of B-Raf proteins. To gain insight into the physiological importance of B-raf alternative splicing, we generated two conditional knockout mice of exons 8b and 9b. Homozygous animals with a constitutive deletion of either exon are healthy and fertile, and survive up to 18 months without any visible abnormalities, demonstrating that alternative splicing is not essential for embryonic development and brain myelination. However, behavioural analyses revealed that expression of exon 9b-containing isoforms is required for B-Raf function in hippocampal-dependent learning and memory. In contrast, mice mutated on exon 8b are not impaired in this function. Interestingly, our results suggest that exon 8b is present only in eutherians and its splicing is differentially regulated among species.

Ablation of BRaf Impairs Neuronal Differentiation in the Postnatal Hippocampus and Cerebellum

“…We further observed that these two bands are absent in tissue extracts from embryos with homozygous exon 3 deletion (lanes del/del; Figure 1B). These genetic data do not support the idea that the known splicing in exons 8b/9b [54] is the basis for the isoforms observed in the immune blots but rather argue in favour of a splicing within exon 3 of the BRaf gene. Sequencing of RT-PCR products confirmed the existence of an intron in exon 3 that was spliced from the mRNA such that in the encoded smaller BRaf protein 33 amino acid residues would be missing.…”

Alternative Splicing in Oncogenic Kinases: From Physiological Functions to Cancer

“…The kinase activity of the different B-Raf isoforms is modulated by the presence or absence of sequences encoded by exon 8b and 9b, located in the hinge region upstream of the kinase domain ( Figure 1 ). While exon 9b is conserved in vertebrates, including fish, amphibians, avians, and mammals, exon 8b appeared later during evolution since it is found only in eutherians, but not in other mammals such as marsupialia and monotremata [ 52 ]. To investigate the physiological relevance of B- raf alternative splicing during development, conditional knockout mice for either exon 8b or exon 9b were generated.…”

Raf Serine/Threonine Protein Kinases: Immunohistochemical Localization in the Mammalian Nervous System

“…Years later, with help of in situ hybridization, it turned out that 24 h following LTP, the B-Raf expression in the dentate gyrus increased significantly (Thomas et al, 1994). Recently, experiments on B-Raf knockout mice proved, that the B-Raf is absolutely necessary for learning and memory consolidation (Chen et al, 2006;Valluet et al, 2010). However, the detailed mechanisms of the participation of B-Raf in LTP are unknown.…”