The quantitative trait locus ahl8 is a key contributor to the early-onset, age-related hearing loss of DBA/2J mice. A nonsynonymous nucleotide substitution in the mouse fascin-2 gene (Fscn2) is responsible for this phenotype, confirmed by wild-type BAC transgene rescue of hearing loss in DBA/2J mice. In chickens and mice, FSCN2 protein is abundant in hair-cell stereocilia, the actin-rich structures comprising the mechanically sensitive hair bundle, and is concentrated toward stereocilia tips of the bundle's longest stereocilia. FSCN2 expression increases when these stereocilia differentially elongate, suggesting that FSCN2 controls filament growth, stiffens exposed stereocilia, or both. Because ahl8 accelerates hearing loss only in the presence of mutant cadherin 23, a component of hair-cell tip links, mechanotransduction and actin crosslinking must be functionally interrelated.

Although CLIC5 is a member of the chloride intracellular channel protein family, its association with actin-based cytoskeletal structures suggests that it may play an important role in their assembly or maintenance. Mice homozygous for a new spontaneous recessive mutation of the Clic5 gene, named jitterbug ( jbg), exhibit impaired hearing and vestibular dysfunction. The jbg mutation is a 97 bp intragenic deletion that causes skipping of exon 5, which creates a translational frame shift and premature stop codon. Western blot and immunohistochemistry results confirmed the predicted absence of CLIC5 protein in tissues of jbg/jbg mutant mice. Histological analysis of mutant inner ears revealed dysmorphic stereocilia and progressive hair cell degeneration. In wild-type mice, CLIC5-specific immunofluorescence was detected in stereocilia of both cochlear and vestibular hair cells and also along the apical surface of Kolliker's organ during cochlear development. Refined immunolocalization in rat and chicken vestibular hair cells showed that CLIC5 is limited to the basal region of the hair bundle, similar to the known location of radixin. Radixin immunostaining appeared reduced in hair bundles of jbg mutant mice. By mass spectrometry and immunoblotting, CLIC5 was shown to be expressed at high levels in stereocilia of the chicken utricle, in an approximate 1:1 molar ratio with radixin. These results suggest that CLIC5 associates with radixin in hair cell stereocilia and may help form or stabilize connections between the plasma membrane and the filamentous actin core.

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During development of the chick cochlea, actin crosslinkers and barbed-end cappers presumably influence growth and remodeling of the actin paracrystal of hair cell stereocilia. We used mass spectrometry to identify and quantify major actin-associated proteins of the cochlear sensory epithelium from E14 to E21, when stereocilia widen and lengthen. Tight actin crosslinkers (i.e. fascins, plastins, and espin) are expressed dynamically during cochlear epithelium development between E7 and E21, with FSCN2 replacing FSCN1 and plastins remaining low in abundance. Capping protein, a barbed-end actin capper, is located at stereocilia tips; it is abundant during growth phase II, when stereocilia have ceased elongating and are increasing in diameter. Capping protein levels then decline during growth phase III, when stereocilia reinitiate barbed-end elongation. Although actin crosslinkers are readily detected by electron microscopy in developing chick cochlea stereocilia, quantitative mass spectrometry of stereocilia isolated from E21 chick cochlea indicated that tight crosslinkers are present there in stoichiometric ratios relative to actin that are much lower than their ratios for vestibular stereocilia. These results demonstrate the value of quantitation of global protein expression in chick cochlea during stereocilia development. Molecular & Cellular Proteomics 13: 10.1074/mcp.M113.033704, 606-620, 2014.The vertebrate hair bundle, an excellent exemplar of intrinsic control of cellular architecture, constitutes 30 -300 actinfilled stereocilia arranged in rows of increasing length but constant diameter. The elegant descriptive studies of the development of bundles from the chick cochlea (basilar papilla) from Lew Tilney and his collaborators established this preparation as ideal for studying stereocilia morphogenesis and bundle assembly (1). They demonstrated that hair cells regulate stereocilia length, width, and number by segregating key steps into distinct phases, which allows the bundle to build itself sequentially; differential regulation and expression of key proteins accordingly must occur during these phases. Because these phases appear to overlap in time during mammalian hair bundle development (2) but are distinct in chick, the chick cochlea is a particularly useful preparation for understanding bundle assembly.Actin filaments in developing chick cochlear stereocilia elongate in three phases (3). In phase I, stereocilia grow to a maximum of 1.5-2.0 m; their actin filaments are initially disordered but crosslinks visible by electron microscopy progressively form and organize filaments into tight parallel bundles (paracrystals). In phase II, elongation is minimal but new actin filaments are added laterally to the stereocilum core, increasing the number of filaments more than 4-fold in apical hair cells. Finally, in phase III, elongation resumes and stereocilia grow to their mature lengths (Figs. 1A, 1B). The discrete nature of these three phases suggests that regulation of elongation and filament number are critic...

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Asymmetric stem cell division has emerged as a major regulatory mechanism for physiologic control of stem cell numbers. Reinvigoration of the cancer stem cell theory suggests that tumorigenesis may be regulated by maintaining the balance between asymmetric and symmetric cell division. Therefore, mutations affecting this balance could result in aberrant expansion of stem cells. Although a number of molecules have been implicated in regulation of asymmetric stem cell division, here, we highlight known tumor suppressors with established roles in this process. While a subset of these tumor suppressors were originally defined in developmental contexts, recent investigations reveal they are also lost or mutated in human cancers. Mutations in tumor suppressors involved in asymmetric stem cell division provide mechanisms by which cancer stem cells can hyperproliferate and offer an intriguing new focus for understanding cancer biology. Our discussion of this emerging research area derives insight from a frontier area of basic science and links these discoveries to human tumorigenesis. This highlights an important new focus for understanding the mechanism underlying expansion of cancer stem cells in driving tumorigenesis.

Purpose: Advances in clinical genomic sequencing capabilities, including reduced costs and knowledge gains, have bolstered the consideration of genomic screening in healthy adult populations. Yet, little is known about the existing landscape of genomic screening programs in the United States. It can be difficult to find information on current implementation efforts and best practices, particularly in light of critical questions about equity, cost, and benefit. Methods: In 2020, we searched publicly available information on the Internet and the scientific literature to identify programs and collect information, including: setting, program funding, targeted population, test offered, and patient cost. Program representatives were contacted throughout 2020 and 2021 to clarify, update, and supplement the publicly available information. Results: Twelve programs were identified. Information was available on key program features, such as setting, genes tested, and target populations. Data on costs, outcomes, or long-term sustainability plans were not always available. Most programs offered testing at no or significantly reduced cost due to generous pilot funding, although the sustainability of these programs remains unknown. Gene testing lists were diverse, ranging from 11 genes (CDC tier 1 genes) to 59 genes (ACMG secondary findings list v.2) to broad exome and genome sequencing. This diversity presents challenges for harmonized data collection and assessment of program outcomes. Conclusions: Early programs are exploring the logistics and utility of population genomic screening in various settings. Coordinated efforts are needed to take advantage of data collected about uptake, infrastructure, and intervention outcomes to inform future research, evaluation, and program development.

An emerging role for DNA sequencing is to identify people at risk for an inherited cancer syndrome in order to prevent or ameliorate the manifestation of symptoms. Two cancer syndromes, Hereditary Breast and Ovarian Cancer and Lynch Syndrome meet the “Tier 1” evidence threshold established by the Centers for Disease Control and Prevention (CDC) for routine testing of patients with a personal or family history of cancer. Advancements in genomic medicine have accelerated public health pilot programs for these highly medically actionable conditions. In this brief report, we provide descriptive statistics from a survey of 746 US respondents from a Qualtrics panel about the public’s awareness of genetic testing, interest in learning about their cancer risk, and likelihood of participating in a population genetic screening (PGS) test. Approximately of half the respondents were aware of genetic testing for inherited cancer risk (n = 377/745, 50.6%) and would choose to learn about their cancer risk (n-309/635, 48.7%). Characteristics of those interested in learning about their cancer risk differed by educational attainment, age, income, insurance status, having a primary care doctor, being aware of genetic testing, and likelihood of sharing information with family (p < 0.05). A sizeable majority of the respondents who were interested in about learning their cancer risk also said that they were likely to participate in a PGS test that involved a clinical appointment and blood draw, but no out-of-pocket cost (n = 255/309, 82.5%). Reasons for not wanting to participate included not finding test results interesting or important, concerns about costs, and feeling afraid to know the results. Overall, our results suggest that engaging and educating the general population about the benefits of learning about an inherited cancer predisposition may be an important strategy to address recruitment barriers to PGS.