Loss of Ahi1 Affects Early Development by Impairing BM88/Cend1-Mediated Neuronal Differentiation

Abstract

Mutations in the Abelson helper integration site-1 (AHI1) gene result in N-terminal Ahi1 fragments and cause Joubert syndrome, an autosomal recessive brain malformation disorder associated with delayed development. How AHI1mutations lead to delayed development remains unclear. Here we report that full-length, but not N-terminal, Ahi1 binds Hap1, a huntingtin-associated protein that is essential for the postnatal survival of mice, and that this binding is regulated during neuronal differentiation by nerve growth factor (NGF). NGF induces dephosphorylation of Hap1A and decreases its association with Ahi1, correlating with increased Hap1A distribution in neurite tips. Consistently, Ahi1 associates with phosphorylated Hap1A in cytosolic, but not in synaptosomal, fractions isolated from mouse brain, suggesting that Ahi1 functions mainly in the soma of neurons. Mass spectrometry analysis of cytosolic Ahi1 immunoprecipitates reveals that Ahi1 also binds Cend1/BM88, a neuronal protein that mediates neuronal differentiation and is highly expressed in postnatal mouse brain. Loss of Ahi1 reduces the levels of Cend1 in the hypothalamus of Ahi1 KO mice, which show retarded growth during postnatal days. Overexpressed Ahi1can stabilize Cend1 in cultured cells. Furthermore, overexpression of Cend1 can rescue the neurite extension defects of hypothalamic neurons from Ahi1 KO mice. Our findings suggest that Cend1 is involved inAhi1-associated hypothalamic neuronal differentiation in early development, giving us fresh insight into the mechanism behind the delayed development in Joubert syndrome.

Cytoplasmic mislocalization of RNA splicing factors and aberrant neuronal gene splicing in TDP-43 transgenic pig brain

“…Cultured cells or brain tissues were homogenized in NP-40,1 × PBS or RIPA buffer with 1× protease inhibitor as previously described [ 57 ]. For immunoprecipitation, the lysates were centrifuged at 800 × g for 5 min, and supernatants were incubated with antibody and Protein A beads (Sigma-Aldrich) at 4 °C overnight, followed by washing with lysis buffer 4 times.…”

“…Culture of primary neurons from mouse brain cortex was performed as described previously [ 57 ]. Briefly, cortical neurons were isolated from the cortex of postnatal day 1 mice.…”

Response to Heller and Bolz

“…Second, Heller and colleagues concluded in their study that the SH3 domain is not essential for AHI1 function based primarily on the finding that two homozygous truncating mutations, Arg1066* and Trp1088Leufs*16, are nonpenetrant in a zebrafish model. This conclusion is slightly overstated because (i) previous studies have reported a functional role of the SH3 domain, 14,15 and the frameshift Trp1088Leufs*16 mutation in the SH3 domain was identified in patients with Joubert syndrome; 8 (ii) the evidence from zebrafish is limited in Heller and colleagues' study 13 because the total number of fish examined is not given, creating a lack of statistical data, and the eyeball size seems to be decreased by e23i23MO injection; and (iii) previous studies have demonstrated that different mutations in the same gene may lead to drastically different retinal phenotypes in mice. 16 We believe that Heller and colleagues' viewpoint is extremely meaningful in this field; however, the exact role of the AHI1 protein domains in a mouse model remains unclear and warrants further investigation.…”

WD40-Repeat Proteins in Ciliopathies and Congenital Disorders of Endocrine System

“…AHI1 mutation is also associated with a broad range of neurological disorders including schizophrenia [107] and autism [108]. Recent mouse model studies have revealed that Ahi1 is highly expressed in the postnatal brain and interacts with other proteins crucial for neuronal differentiation [109,110]. Table 3 lists the WDR proteins involved in endocrine disorders, many of which are often presented as a part of a ciliopathy.…”