GATA3 germline variant is associated with CRLF2 expression and predicts outcome in pediatric B‐cell precursor acute lymphoblastic leukemia

The germline variant at rs3824662 in GATA3 is a risk locus for Philadelphia‐like acute lymphoblastic leukemia (Ph‐like ALL), the biological subtype of B‐cell precursor (BCP)‐ALL defined by a distinct gene expression profile and the presence of specific somatic aberrations including rearrangements of CRLF2. In this study, we investigated whether rs3824662 in GATA3 associates with CRLF2 expression in leukemic cells and predicts prognosis in pediatric BCP‐ALL patients treated according to the ALL Intercontinental Berlin‐Frankfurt‐Münster (IC BFM) 2009 (n = 645) and the ALL IC BFM 2002 (n = 216) protocols. High expression of CRLF2 was observed at both protein and mRNA levels (fourfold higher in AA than in CA + CC) among GATA3 AA variant carriers, independent of the presence of P2RY8‐CRLF2 fusion. Additionally, the AA variant at rs3824662 was a significant factor affecting minimal residual disease level at the end of induction phase and overall survival regardless of the risk group and the protocol. The germline variant at rs3824662 in GATA3 is a prognostic factor which associates with CRLF2 expression in leukemic cells supporting the hypothesis that GATA3 may have a regulatory effect on the CRLF2 pathway in pediatric BCP‐ALL.     

Architecture of the mouse utricle: macular organization and hair bundle heights

Hair bundles are critical to mechanotransduction by vestibular hair cells, but quantitative data are lacking on vestibular bundles in mice or other mammals. Here we quantify bundle heights and their variation with macular locus and hair cell type in adult mouse utricular macula. We also determined that macular organization differs from previous reports. The utricle has approximately 3,600 hair cells, half on each side of the line of polarity reversal (LPR). A band of low hair cell density corresponds to a band of calretinin-positive calyces, i.e., the striola. The relation between the LPR and the striola differs from previous reports in two ways. First, the LPR lies lateral to the striola instead of bisecting it. Second, the LPR follows the striolar trajectory anteriorly, but posteriorly it veers from the edge of the striola to reach the posterior margin of the macula. Consequently, more utricular bundles are oriented mediolaterally than previously supposed. Three hair cell classes are distinguished in calretinin-stained material: type II hair cells, type ID hair cells contacting calretinin-negative (dimorphic) afferents, and type IC hair cells contacting calretinin-positive (calyceal) afferents. They differ significantly on most bundle measures. Type II bundles have short stereocilia. Type IC bundles have kinocilia and stereocilia of similar heights, i.e., KS ratios (ratio of kinocilium to stereocilia heights) approximately 1, unlike other receptor classes. In contrast to these class-specific differences, bundles show little regional variation except that KS ratios are lowest in the striola. These low KS ratios suggest that bundle stiffness is greater in the striola than in the extrastriola.     

ABLIM1 splicing is abnormal in skeletal muscle of patients with DM1 and regulated by MBNL,CELF and PTBP1

Myotonic dystrophy type 1 (DM1) is an RNA‐mediated disorder characterized by muscle weakness, cardiac defects and multiple symptoms and is caused by expanded CTG repeats within the 3′ untranslated region of the DMPK gene. In this study, we found abnormal splicing of actin‐binding LIM protein 1 (ABLIM1) in skeletal muscles of patients with DM1 and a DM1 mouse model (HSA^LR). An exon 11 inclusion isoform is expressed in skeletal muscle and heart of non‐DM1 individuals, but not in skeletal muscle of patients with DM1 or other adult human tissues. Moreover, we determined that ABLIM1 splicing is regulated by several splice factors, including MBNL family proteins, CELF1, 2 and 6, and PTBP1, using a cellular splicing assay. MBNL proteins promoted the inclusion of ABLIM1 exon 11, but other proteins and expanded CUG repeats repressed exon 11 of ABLIM1. This result is consistent with the hypothesis that MBNL proteins are trapped by expanded CUG repeats and inactivated in DM1 and that CELF1 is activated in DM1. However, activation of PTBP1 has not been reported in DM1. Our results suggest that the exon 11 inclusion isoform of ABLIM1 may have a muscle‐specific function, and its abnormal splicing could be related to muscle symptoms of DM1.     

GATA-binding protein-3 regulates T helper type 2 cytokine and ifng loci through interaction with metastasis-associated protein 2

GATA‐binding protein‐3 (GATA‐3) regulates the T helper type 2 (Th2) cytokine locus through induction of chromatin remodelling. However, the molecular mechanism for this is poorly understood. To understand this mechanism better, we screened GATA‐3 interacting proteins using affinity purification and mass spectrometry. We found that GATA‐3 bound to metastasis‐associated protein 2 (MTA‐2), a component of the NuRD chromatin remodelling complex. GATA‐3 and MTA‐2 in turn bound to several regulatory regions of the Th2 cytokine locus and the ifng promoter. Cell transfection assay showed that MTA‐2 acted as an antagonist with GATA‐3 in the expression of Th2 cytokines, but co‐operated with GATA‐3 in the repression of the ifng gene expression. These results suggest that GATA‐3 interacts with MTA‐2 to co‐ordinately regulate Th2 cytokine and ifng loci during T helper cell differentiation.     

V(D)J targeting mistakes occur at low frequency in acute lymphoblastic leukemia

Translocations of proto‐oncogenes to the B‐cell or T‐cell antigen receptor loci in acute T‐ or B‐cell leukemia and lymphoma have been, in most cases, accredited to V(D)J or switch recombination depending on the location of the breakpoint at the receptor locus. Only in rare instances, the reports take into account mechanistic characteristics of the translocation mechanism. To assess the functional ability of several sites implicated in supposedly V(D)J‐mediated translocations, we tested five sites at four proto‐oncogene loci in an ex vivo recombination substrate assay for their potential to act as direct target for V(D)J recombination. Our results show that the LMO2/RBTN2/TTG2 site and one LCK/P56 site readily engage in recombination with a genuine TCR element with the majority of breakpoint junctions showing the characteristics of V(D)J recombination, which strongly supports the involvement of this mechanism in the pathogenesis of the corresponding translocations in vivo. The site at the TLX1/HOX11 locus yielded 0.8% V(D)J‐specific junctions. Sites at the LCK/P56 and TCF3/E2A proto‐oncogenes resulted in exclusively unspecific breakpoints scattered over part of or the entire proto‐oncogene region tested, marking them as unlikely V(D)J recombination targets. Our data suggest that, while being a potentially dangerous mechanism due to the introduction of DNA breaks, V(D)J recombination is a tightly controlled mechanism allowing for only few direct mistakes. © 2009 Wiley‐Liss, Inc.     

LMO2 promotes angiogenesis probably by up-regulation of bFGF in endothelial cells: an implication of its pathophysiological role in infantile haemangioma

Sun Z‐J, Cai Y, Chen G, Wang R, Jia J, Chen X‐M, Zheng L‐W & Zhao Y‐F
(2010) Histopathology 57 , 622–632
LMO2 promotes angiogenesis probably by up‐regulation of bFGF in endothelial cells: an implication of its pathophysiological role in infantile haemangioma Aims: Infantile haemangiomas (IHs) are common benign vascular tumours distinctive for their perinatal presentation, rapid growth during the first year of life and subsequent slow involution. Recent research has indicated that endothelial cells of haemangiomas express LIM‐only protein 2 (LMO2). The aim of this study was to investigate the role of LMO2 in the pathogenesis of IHs was investigated. Methods and results: Immunoreactivity of LMO2 was assessed in specimens of 19 IH. Stable transfection of LMO2 into human endothelial cell lines (EAhy926) was performed to evaluate the role of LMO2 in terms of the change in cell proliferation, cell cycle and cell migration as well as the expression level of angiogenic factors. Immunoreactivity for LMO2 was detected in all IH specimens, specifically in the nucleus of the endothelial cells. The intensity of LMO2 immunostaining decreased significantly from proliferative to involuting stages. Furthermore, the overexpression of LMO2 enhanced the proliferation and migration of the endothelial cells and promoted G0/G1–S‐phase transition in vitro, together with an up‐regulation of bFGF expression. Conclusions: LMO2 probably promotes angiogenesis by up‐regulation of bFGF expression and thereby consequently influences progression of IH.     

Interspecific Variations of Inner Ear Structure in the Deep‐Sea Fish Family Melamphaidae

Inner ear structures are compared among three major genera of the deep‐sea fish family Melamphaidae (bigscales and ridgeheads). Substantial interspecific variation is found in the saccular otoliths, including the presence of a unique otolithic “spur” in the genera Melamphaes and Poromitra. The variation in the saccular otolith is correlated with an increase in the number of hair bundle orientation groups on the sensory epithelia from the genera Scopelogadus to Poromitra to Melamphaes. The diverse structural variations found in the saccule may reflect the evolutionary history of these species. The sensory hair cell bundles in this family have the most variable shapes yet encountered in fish ears. In the saccule, most of the hair bundles are 15–20 μm high, an exceptional height for fish otolithic end organs. These bundles have large numbers of stereovilli, including some that reach the length of the kinocilium. In the utricle, the striolar region separates into two unusually shaped areas that have not been described in any other vertebrates. The brains in all species have a relatively small olfactory bulb and optic tectum, as well as an enlarged posterior cerebellar region that is likely to be involved in inner ear and lateral line (octavolateral) functions. Data from melamphaids support the hypothesis that specialized anatomical structures are found in the ears of some (if not most) deep‐sea fishes, presumably enhancing their hearing sensitivity. Anat Rec, 296:1064–1082, 2013. © 2013 Wiley Periodicals, Inc.     

Muscleblind‐like 1 is a negative regulator of TGF‐β‐dependent epithelial–mesenchymal transition of atrioventricular canal endocardial cells

The development of the valves and septa of the heart depends on the formation and remodeling of endocardial cushions. Here, we report that the alternative splicing regulator muscleblind‐like 1 (MBNL1) exhibits a regionally restricted pattern of expression in canal region endocardium and ventricular myocardium during endocardial cushion development in chicken. Knockdown of MBNL1 in atrioventricular explants leads to a transforming growth factor β‐dependent increase in epithelial–mesenchymal transition (EMT) of endocardial cells. This reveals a novel role for MBNL1 during embryonic development, and represents the first evidence that an alternative splicing regulator is a key player in endocardial cushion development. Developmental Dynamics 238:3266–3272, 2009. © 2009 Wiley‐Liss, Inc.     

Spatial location and strength of BOLD activation in high-spatial-resolution fMRI of the motor cortex: a comparison of spin echo and gradient echo fMRI at 7 T

The increased blood oxygenation level‐dependent contrast‐to‐noise ratio at ultrahigh field (7 T) has been exploited in a comparison of the spatial location and strength of activation in high‐resolution (1.5 mm isotropic) gradient echo (GE) and spin echo (SE), echo planar imaging data acquired during the execution of a simple motor task in five subjects. SE data were acquired at six echo times from 30 to 55 ms. Excellent fat suppression was achieved in the SE echo planar images using slice‐selective gradient reversal. Threshold‐free cluster enhancement was used to define regions of interest (ROIs) containing voxels showing significant stimulus‐locked signal changes from the GE and average SE data. These were used to compare the signal changes and spatial locations of activated regions in SE and GE data. T₂ and T₂* values were measured, with means of 48.3 ± 1.1 ms and 36.5 ± 3.4 ms in the SE ROI. In addition, we identified a dark band in SE images of the motor cortex corresponding to a region in which T₂ and T₂* were significantly lower than in the surrounding grey matter. The fractional SE signal change in the ROI was found to vary linearly as a function of TE, with a slope that was dependent on the particular ROI assessed: the mean ΔR₂ value was found to be 0.85 ± 0.11 s^–1 for the SE ROI and ?0.37 ± 0.05 s^–1 for the GE ROI. The fractional signal change relative to the shortest TE revealed that the largest signal change occurred at a TE of 45 ms outside of the dark band. At this TE, the ratio of the fractional signal change in GE and SE data was found to be 0.48 ± 0.05. Phase maps produced from high‐resolution GE images spanning the right motor cortex were used to identify veins. The GE ROI was found to contain 18% more voxels overlying the venous mask than the SE ROI. Copyright © 2011 John Wiley & Sons, Ltd.     

The vestibular nerve of the chinchilla. III. Peripheral innervation patterns in the utricular macula

1. Nerve fibers supplying the utricular macula of the chinchilla were labeled by extracellular injection of horseradish peroxidase into the vestibular nerve. The peripheral terminations of individual fibers were reconstructed and related to the regions of the end organ they innervated and to the sizes of their parent axons. 2. The macula is divided into medial and lateral parts by the striola, a narrow zone that runs for almost the entire length of the sensory epithelium. The striola can be distinguished from the extrastriolar regions to either side of it by the wider spacing of its hair cells. Calyx endings in the striola have especially thick walls, and, unlike similar endings in the extrastriola, many of them innervate more than one hair cell. The striola occupies 10% of the sensory epithelium; the lateral extrastriola, 50%; and the medial extrastriola, 40%. 3. The utricular nerve penetrates the bony labyrinth anterior to the end organ. Axons reaching the anterior part of the sensory epithelium run directly through the connective tissue stroma. Those supplying more posterior regions first enter a fiber layer located at the bottom of the stroma. Approximately one-third of the axons bifurcate below the epithelium, usually within 5-20 microns of the basement membrane. Bifurcations are more common in fibers destined for the extrastriola than for the striola. 4. Both calyx and bouton endings were labeled. Calyces can be simple or complex. Simple calyces innervate individual hair cells, whereas complex calyces supply 2-4 adjacent hair cells. Complex endings are more heavily concentrated in the striola than in the extrastriola. Simple calyces and boutons are found in all parts of the epithelium. Calyces emerge from the parent axon or one of its thick branches. Boutons, whether en passant or terminal, are located on thin collaterals. 5. Fibers can be classified into calyx, bouton, or dimorphic categories. The first type only has calyx endings; the second, only bouton endings; and the third, both kinds of endings. Calyx units make up 6% of the labeled fibers, bouton units less than 2%, and dimorphic units greater than 92%. The three fiber types differ in the macular zones they supply and in the diameters of their parent axons. Calyx units were restricted to the striola. The few bouton units were found in the extrastriola.(ABSTRACT TRUNCATED AT 400 WORDS)     

The transcription factor D‐Pax2 regulates crystallin production during eye development in Drosophila melanogaster

The generation of a functioning Drosophila eye requires the coordinated differentiation of multiple cell types and the morphogenesis of eye‐specific structures. Here we show that D‐Pax2 plays a significant role in lens development through regulation of the Crystallin gene and because Crystallin is also expressed in D‐Pax2⁺ cells in the external sensory organs. Loss of D‐Pax2 function leads to loss of Crystallin expression in both eyes and bristles. A 2.3 kilobase (kb) upstream region of the Crystallin gene can drive GFP expression in the eye and is dependent on D‐Pax2. In addition, D‐Pax2 binds to an evolutionarily conserved site in this region that, by itself, is sufficient to drive GFP expression in the eye. However, mutation of this site does not greatly affect the regulatory region's function. The data indicate that D‐Pax2 acts to promote lens development by controlling the production of the major protein component of the lens. Whether this control is direct or indirect remains unresolved. Developmental Dynamics 238:2530–2539, 2009. © 2009 Wiley‐Liss, Inc.     

Regulation of the Caenorhabditis elegans posterior hox gene egl‐5 by microRNA and the polycomb‐like gene sop‐2

In Caenorhabditis elegans, the domains of Hox gene expression are controlled by the novel global regulatory gene sop‐2. We identified a region located 3′ of the Hox gene egl‐5 that promotes ectopic expression of an egl‐5 reporter gene in a sop‐2 mutant. SOP‐2 could directly block positive regulatory factors acting in this region, or it could block their expression. We identified three possible miRNA binding sites within the egl‐5 3′ untranslated region (UTR). Cognate microRNAs are expressed in relevant tissues and can block egl‐5 expression when expressed from a transgene. Mutation of the putative binding sites in the egl‐5 3′UTR resulted in a modest degree of misexpression of a minimal egl‐5 reporter gene, suggesting that microRNAs may contribute to the tight restriction of egl‐5 expression to particular cell lineages. Developmental Dynamics 238:595–603, 2009. © 2009 Wiley‐Liss, Inc.