Primary ciliary dyskinesia (PCD) is a genetically heterogeneous autosomal recessive disorder characterized by recurrent infections of the respiratory tract associated with abnormal function of motile cilia. Approximately half of PCD patients also have alterations in the left-right organization of internal organ positioning including situs inversus and situs ambiguous (Kartagener’s Syndrome, KS). Here we identify an uncharacterized coiled-coil domain containing protein (CCDC40) essential for correct left-right patterning in mouse, zebrafish and humans. Ccdc40 is expressed in tissues that contain motile cilia and mutation of Ccdc40 results in cilia with reduced ranges of motility. Importantly, we demonstrate that CCDC40 deficiency causes a novel PCD variant characterized by misplacement of central pair microtubules and defective axonemal assembly of inner dynein arms (IDAs) and dynein regulator complexes (DRCs). CCDC40 localizes to motile cilia and the apical cytoplasm and is responsible for axonemal recruitment of CCDC39, which is also mutated in a similar PCD variant.

SUMMARY During development of the embryonic neocortex, tightly regulated expansion of neural stem cells (NSCs) and their transition to intermediate progenitors (IPs) are critical for normal cortical formation and function. Molecular mechanisms that regulate NSC expansion and transition remain unclear. Here, we demonstrate that the microRNA (miRNA) miR-17-92 cluster is required for maintaining proper populations of cortical radial glial cells (RGCs) and IPs through repression of Pten and Tbr2 protein. Knockout of miR-17-92 and its paralogs specifically in the developing neocortex restricts NSC proliferation, suppresses RGC expansion, and promotes transition of RGCs to IPs. Moreover, Pten and Tbr2 protectors specifically block silencing activities of endogenous miR-17-92 and control proper numbers of RGCs and IPs in vivo. Our results demonstrate a critical role for miRNAs in promoting NSC proliferation and modulating the cell-fate decision of generating distinct neural progenitors in the developing neocortex.

IntroductionHereditary hemochromatosis is a common disorder in humans, characterized by iron overload resulting in tissue injury and ultimately organ failure. Typically, hemochromatosis exhibits an autosomal-recessive pattern of inheritance and is associated with mutations in HFE, hemojuvelin, hepcidin, or transferrin receptor 2. 1,2 Targeted deletion of these genes in the mouse results in hemochromatosis, providing mouse models for most forms of the disease. Hemochromatosis type IV, also referred to as ferroportin (Fpn) disease, results from mutations in the iron transporter ferroportin. Fpn is the only known iron exporter in mammalian cells and is present on the surface of macrophages, intestinal enterocytes, hepatocytes, and placental cells. [3][4][5] The level of cell surface Fpn is regulated by its interaction with hepcidin, a peptide secreted by the liver in response to iron stores and inflammation. Hepcidin binds to Fpn, inducing its internalization and degradation, thus regulating the export of iron from cells to plasma. 6 Mutations in Fpn lead to iron-overload disease but, in contrast to other forms of hemochromatasis, ferroportin disease exhibits an autosomal-dominant pattern of inheritance. 7 The disorder has different presentations depending on the Fpn mutation. Mutations leading to Fpn that is not internalized by hepcidin result in iron accumulation in hepatocytes and high transferrin saturation. 8,9 Mutations leading to Fpn that is not appropriately targeted to the cell surface result in iron accumulation in Kupffer cells and low transferrin saturation. [9][10][11] The mechanism by which the disease mutations exert a dominant effect is unclear. Some groups that study the disease suggest that it results from haploinsufficiency, 10,12 whereas others suggest that the disorder results from a dominantnegative effect of the mutant allele. 9,13 Importantly, all human mutations are missense mutations and mice that are heterozygous for a targeted deletion of Fpn do not show the disease. 14 Treatment for hemochromatosis aims to decrease iron load by repeated phlebotomy and this treatment works well for most patients. Many patients with ferroportin disease, however, become anemic with phlebotomy, highlighting the need for a mouse model to develop better treatments.We report here on a missense mutation in mouse Fpn that results in a disorder that is identical to classic human ferroportin disease. We show that macrophages isolated from mutant mice have no Fpn on their cell surface and that expression of Fpn constructs containing the missense mutation (H32R) affects the behavior of wild-type Fpn. These results show that Fpn disease is due to a dominant-negative effect of the mutant allele and provide the first mouse model for this disorder. Materials and methods Generation of mutant mice and identification of Fpn mutationThe ffe mouse line was identified in a screen for recessive ethylnitrosourea (ENU)-induced mutations that cause morphologic abnormalities at embryonic day (E) 12.5. [15][16][17] The ffe mutation...

The precise organization of motor neuron subtypes in a columnar pattern in developing spinal cords is controlled by cross-interactions of multiple transcription factors and segmental expressions of Hox genes and their accessory proteins. Accurate expression levels and domains of these regulators are essential for organizing spinal motor neuron columns and axonal projections to target muscles. Here, we show that microRNA miR-9 is transiently expressed in a motor neuron subtype and displays overlapping expression with its target gene FoxP1. Overexpression or knockdown of miR-9 alters motor neuron subtypes, switches columnar identities, and changes axonal innervations in developing chick spinal cords. miR-9 modifies spinal columnar organization by specifically regulating FoxP1 protein levels, which in turn determine distinct motor neuron subtypes. Our findings demonstrate that miR-9 is an essential regulator of motor neuron specification and columnar formation. Moreover, the overlapping expression of miR-9 and its target FoxP1 further illuminates the importance of fine-tuning regulation by microRNAs in motor neuron development.