Maternally derived 15q11.2-q13.1 duplication and H19-DMR hypomethylation in a patient with Silver?Russell syndrome

Silver?Russell syndrome (SRS) is a congenital developmental disorder characterized by intrauterine and postnatal growth failure, craniofacial features (including a triangular shaped face and broad forehead), relative macrocephaly, protruding forehead, body asymmetry and feeding difficulties. Hypomethylation of the H19 differentially methylated region (DMR) on chromosome 11p15.5 is the most common cause of the SRS phenotype. We report the first SRS patient with hypomethylation of the H19-DMR and maternally derived 15q11.2-q13.1 duplication. Although her clinical manifestations overlapped with those of previously reported SRS cases, the patient’s intellectual disability and facial dysmorphic features were inconsistent with the SRS phenotype. Methylation analyses, array comparative genomic hybridization, and a FISH analysis revealed the hypomethylation of the H19-DMR and a maternally derived interstitial 5.7?Mb duplication at 15q11.2-q13.1 encompassing the Prader?Willi/Angelman critical region in the patient. On the basis of the genetic and clinical findings in the present and previously reported cases, it is unlikely that the 15q duplication in the patient led to the development of hypomethylation of the H19-DMR and it is reasonable to consider that the characteristic phenotype in the patient was caused by the coexistence of the two (epi)genetic conditions. Further studies are needed to clarify the mechanisms leading to methylation aberrations in SRS

Ran and Calcineurin Can Participate Collaboratively in the Regulation of Spermatogenesis in Scallop

Calcineurin is a calcium/calmodulin-dependent protein phosphatase that plays important roles in the transduction of calcium signals in a variety of tissues. In addition, calcineurin has been implicated in the process of spermatogenesis. A novel calcineurin-binding protein, CaNBP75, has been identified in scallop testis. The C-terminal region of CaNBP75 is homologous to the C-terminal region of RanBP3, a Ran binding domain-containing protein. A small G-protein Ran has been involved in spermiogenesis by virtue of the fact that its localization in spermatids changes during spermiogenesis. The current study was performed to investigate the functions of Ran and CaNBP75 in the regulation of calcineurin in testis to further understand the basic functions of calcineurin during spermatogenesis. First, cloning and sequencing of a scallop Ran cDNA isolated from testis revealed that scallop Ran is well-conserved at the amino acid level. Secondly, direct binding of Ran to CaNBP75 was demonstrated in an in vitro pull-down assay. Thirdly, analysis of the tissue distribution of Ran, CaNBP75 and calcineurin showed that these proteins are abundantly expressed in testis. Fourthly, comparison of the expression profiles of Ran and CaNBP75 with that of calcineurin in scallop testis during the maturation cycle revealed that Ran and CaNBP75 mRNA levels increase during meiosis and spermiogenesis, similar to calcineurin. Finally, co-immunoprecipitation analysis suggests that Ran, CaNBP75 and calcineurin interact in scallop testis during maturation. These results suggest that Ran, CaNBP75, and calcineurin may act in a coordinated manner to regulate spermatogenesis

Reconstituted Human Myosin Light Chain Phosphatase Reveals Distinct Roles of Two Inhibitory Phosphorylation Sites of the Regulatory Subunit, MYPT1

The myosin light chain phosphatase (MLCP) is a cytoskeleton-associated protein phosphatase-1 (PP1) holoenzyme and a RhoA/ROCK effector, regulating cytoskeletal reorganization. ROCK-induced phosphorylation of the MLCP regulatory subunit (MYPT1) at two sites, Thr696 and Thr853, suppresses the activity, although little is known about the difference in the role. Here, we developed a new method for the preparation of the recombinant human MLCP complex and determined the molecular and cellular basis of inhibitory phosphorylation. The recombinant MLCP partially purified from mammalian cell lysates retained characteristics of the native enzyme, such that it was fully active without Mn²⁺ and sensitive to PP1 inhibitor compounds. Selective thio-phosphorylation of MYPT1 at Thr696 with ROCK inhibited the MLCP activity 30%, whereas the Thr853 thio-phosphorylation did not alter the phosphatase activity. Interference with the docking of phospho-Thr696 at the active site weakened the inhibition, suggesting selective autoinhibition induced by phospho-Thr696. Both Thr696 and Thr853 sites underwent autodephosphorylation. Compared with that of Thr853, phosphorylation of Thr696 was more stable, and it facilitated Thr853 phosphorylation. Endogenous MYPT1 at Thr696 was spontaneously phosphorylated in quiescent human leiomyosarcoma cells. Serum stimulation of the cells resulted in dissociation of MYPT1 from myosin and PP1C in parallel with an increase in the level of Thr853 phosphorylation. The C-terminal domain of human MYPT1(495–1030) was responsible for the binding to the N-terminal portion of myosin light meromyosin. The spontaneous phosphorylation at Thr696 may adjust the basal activity of cellular MLCP and affect the temporal phosphorylation at Thr853 that is synchronized with myosin targeting

Partial internal biliary diversion for patients with progressive familial intrahepatic cholestasis type 1.

We herein report a case of progressive familial intrahepatic cholestasis with partial internal biliary diversion (PIBD). Although by using PIBD an external stoma can be avoided, exposure of the ileocecal junction to bile reflux as well as the effects of the direct bile flow on the colonic mucosa require further investigation

Identification of a novel heterozygous mutation of the Aggrecan gene in a family with idiopathic short stature and multiple intervertebral disc herniation

Aggrecan is a critical proteoglycan component of the extracellular matrix of the growth plates and articular cartilage and has a key role in the biophysical and biomechanical properties of cartilage. Recently, heterozygous mutations in the ACAN gene, which encodes aggrecan, have been identified in patients with short stature and accelerated bone age. We herein report another family with a heterozygous ACAN mutation associated with idiopathic short stature along with accelerated bone age and early-onset herniation of the lumbar discs at the levels of L1/2 through L5/S1. Whole-exome sequencing identified a novel heterozygous frameshift mutation in the ACAN gene (c.1744delT; p.Phe582fs*69) in all of the affected family members but not in the unaffected one, providing further evidence that ACAN haploinsufficiency causes short stature with advanced bone maturation. In addition, we advocate early-onset multiple disc herniation as a novel phenotype associated with ACAN haploinsufficiency

Newborn screening for mucopolysaccharidoses: a pilot study of measurement of glycosaminoglycans by tandem mass spectrometry

Background Mucopolysaccharidoses (MPS) are a group of inborn errors of metabolism that are progressive and usually result in irreversible skeletal, visceral, and/or brain damage, highlighting a need for early diagnosis. Methods This pilot study analyzed 2862 dried blood spots (DBS) from newborns and 14 DBS from newborn patients with MPS ( MPS I, n = 7; MPS II, n = 2; MPS III, n = 5). Disaccharides were produced from polymer GAGs by digestion with chondroitinase B, heparitinase, and keratanase II. Heparan sulfate (0S, NS), dermatan sulfate (DS) and mono-and di-sulfated KS were measured by liquid chromatography tandem mass spectrometry (LC-MS/MS). Median absolute deviation (MAD) was used to determine cutoffs to distinguish patients from controls. Cutoffs were defined as median + 7x MAD from general newborns. Results The cutoffs were as follows: HS-0S > 90 ng/mL; HS-NS > 23 ng/mL, DS > 88 ng/mL; mono-sulfated KS > 445 ng/mL; di-sulfated KS > 89 ng/mL and ratio di-KS in total KS > 32 %. All MPS I and II samples were above the cutoffs for HS-0S, HS-NS, and DS, and all MPS III samples were above cutoffs for HS-0S and HS-NS. The rate of false positives for MPS I and II was 0.03 % based on a combination of HS-0S, HSNS, and DS, and for MPS III was 0.9 % based upon a combination of HS-0S and HS-NS. Conclusions Combination of levels of two or more different GAGs improves separation of MPS patients from unaffected controls, indicating that GAG measurements are potentially valuable biomarkers for newborn screening for MP