Chromosomal Dynamics at the Shh Locus: Limb Bud-Specific Differential Regulation of Competence and Active Transcription

The expression of Sonic hedgehog (Shh) in mouse limb buds is regulated by a long-range enhancer 1 Mb upstream of the Shh promoter. We used 3D-FISH and chromosome conformation capture assays to track changes at the Shh locus and found that long-range promoter-enhancer interactions are specific to limb bud tissues competent to express Shh. However, the Shh locus loops out from its chromosome territory only in the posterior limb bud (zone of polarizing activity or ZPA), where Shh expression is active. Notably, while Shh mRNA is detected throughout the ZPA, enhancer-promoter interactions and looping out were only observed in small fractions of ZPA cells. In situ detection of nascent Shh transcripts and unstable EGFP reporters revealed that active Shh transcription is likewise only seen in a small fraction of ZPA cells. These results suggest that chromosome conformation dynamics at the Shh locus allow transient pulses of Shh transcription

Chromosomal Dynamics at the Shh Locus: Limb Bud-Specific Differential Regulation of Competence and Active Transcription

SummaryThe expression of Sonic hedgehog (Shh) in mouse limb buds is regulated by a long-range enhancer 1 Mb upstream of the Shh promoter. We used 3D-FISH and chromosome conformation capture assays to track changes at the Shh locus and found that long-range promoter-enhancer interactions are specific to limb bud tissues competent to express Shh. However, the Shh locus loops out from its chromosome territory only in the posterior limb bud (zone of polarizing activity or ZPA), where Shh expression is active. Notably, while Shh mRNA is detected throughout the ZPA, enhancer-promoter interactions and looping out were only observed in small fractions of ZPA cells. In situ detection of nascent Shh transcripts and unstable EGFP reporters revealed that active Shh transcription is likewise only seen in a small fraction of ZPA cells. These results suggest that chromosome conformation dynamics at the Shh locus allow transient pulses of Shh transcription

Efficacy of erlotinib and imatinib in a patient with a rectal gastrointestinal stromal tumor and synchronous pulmonary adenocarcinoma : A case report

The synchronous existence of lung cancer and gastrointestinal stromal tumors (GIST) is considered to be extremely rare. To the best of our knowledge, this is the first report about the treatment of lung cancer and GIST with two kinds of molecular targeting drugs. An 83-year-old woman with a rectal GIST, which carried a c-kit mutation, and pulmonary adenocarcinoma, which exhibited an epidermal growth factor receptor (EGFR) mutation, was treated alternately with imatinib and erlotinib. Good control over both diseases was achieved for two years. The present case is not only of interest due to the rare co-occurrence of GIST and lung cancer, but also because it involved two tumors carrying different gene mutations, and both tumors were brought under control using different molecular targeting drugs

Utility of Nd isotope ratio as a tracer of marine animals : regional variation in coastal seas and causal factors

Isotopic compositions of animal tissue are an intrinsic marker commonly used to trace animal origins and migrations; however, few isotopes are effective for this purpose in marine environments, especially on a local scale. The isotope ratio of the lanthanoid element neodymium (Nd) is a promising tracer for coastal animal migrations. Neodymium derives from the same geologic materials as strontium, well known as an isotopic tracer (87Sr/86Sr) for terrestrial and anadromous animals. The advantage of the Nd isotope ratio (143Nd/144Nd, expressed as εNd) is that it varies greatly in the ocean according to the geology of the neighboring continents, whereas oceanic 87Sr/86Sr is highly uniform. This study explored the utility of the Nd isotope ratio as a marine tracer by investigating the variation of εNd preserved in tissues of coastal species, and the causes of that variation, in a region of northeastern Japan where the bedrock geology is highly variable. We measured εNd and 87Sr/86Sr in seawater, river water, and soft tissues of sedentary suspension feeders: the mussels Mytilus galloprovincialis and Mytilus coruscus and the oyster Crassostrea gigas. We also measured concentrations of three lanthanoids (La, Ce, and Pr) in shellfish bodies to determine whether the Nd in shellfish tissue was derived from solution in seawater or from suspended particulates. The εNd values in shellfish tissue varied regionally (−6 to +1), matching the ambient seawater, whereas all 87Sr/86Sr values were homogeneous and typical of seawater (0.7091–0.7092). The seawater εNd values were in turn correlated with those in the adjacent rivers, linking shellfish εNd to the geology of river catchments. The depletion of Ce compared to La and Pr (negative Ce anomaly) suggested that the Nd in shellfish was derived from the dissolved phase in seawater. Our results indicate that the distinct Nd isotope ratio derived from local geology is imprinted, through seawater, on the soft tissues of shellfish. This result underscores the potential of εNd as a tracer of coastal marine animals

Two Types of Etiological Mutation in the Limb-Specific Enhancer of Shh

An enhancer named MFCS1 regulates Sonic hedgehog (Shh) expression in the posterior mesenchyme of limb buds. Several mutations in MFCS1 induce ectopic Shh expression in the anterior limb bud, and these result in preaxial polydactyly (PPD). However, the molecular basis of ectopic Shh expression remains elusive, although some mutations are known to disrupt the negative regulation of Shh expression in the anterior limb bud. Here, we analyzed the molecular mechanism of ectopic Shh expression in PPD including in a mouse mutation—hemimelic extra toes (Hx)—and in other MFCS1 mutations in different species. First, we generated transgenic mouse lines with a LacZ reporter cassette flanked with tandem repeats of 40 bp MFCS1 fragments harboring a mutation. The transgenic mouse line with the Hx-type fragment showed reporter expression exclusively in the anterior, but not in the posterior margins of limb buds. In contrast, no specific LacZ expression was observed in lines carrying the MFCS1 fragment with other mutations. Yeast one-hybrid assays revealed that the msh-like homeodomain protein, MSX1, bound specifically to the Hx sequence of MFCS1. Thus, PPD caused by mutations in MFCS1 has two major types of molecular etiology: loss of a cis-motif for negative regulation of Shh, and acquisition of a new cis-motif binding to a preexisting transcription factor, as represented by the Hx mutation

Two Types of Etiological Mutation in the Limb-Specific Enhancer of Shh

An enhancer named MFCS1 regulates Sonic hedgehog (Shh) expression in the posterior mesenchyme of limb buds. Several mutations in MFCS1 induce ectopic Shh expression in the anterior limb bud, and these result in preaxial polydactyly (PPD). However, the molecular basis of ectopic Shh expression remains elusive, although some mutations are known to disrupt the negative regulation of Shh expression in the anterior limb bud. Here, we analyzed the molecular mechanism of ectopic Shh expression in PPD including in a mouse mutation—hemimelic extra toes (Hx)—and in other MFCS1 mutations in different species. First, we generated transgenic mouse lines with a LacZ reporter cassette flanked with tandem repeats of 40 bp MFCS1 fragments harboring a mutation. The transgenic mouse line with the Hx-type fragment showed reporter expression exclusively in the anterior, but not in the posterior margins of limb buds. In contrast, no specific LacZ expression was observed in lines carrying the MFCS1 fragment with other mutations. Yeast one-hybrid assays revealed that the msh-like homeodomain protein, MSX1, bound specifically to the Hx sequence of MFCS1. Thus, PPD caused by mutations in MFCS1 has two major types of molecular etiology: loss of a cis-motif for negative regulation of Shh, and acquisition of a new cis-motif binding to a preexisting transcription factor, as represented by the Hx mutation