Structural Protein Analysis of Driver Gene Mutations in Conjunctival Melanoma.

In recent years, there has been tremendous enthusiasm with respect to detailing the genetic basis of many neoplasms, including conjunctival melanoma (CM). We aim to analyze five proteins associated with CM, namely BRAF, NRAS, c-KIT, NF1, and PTEN. We evaluated each protein for its intrinsically disordered protein regions (IDPRs) and its protein-protein interactions (PPI) with the Predictor of Natural Disordered Protein Regions (PONDR®) and the Search Tool for the Retrieval of Interacting Genes (STRING®). Our PONDR® analysis found high levels of IDPRs in all five proteins with mutations linked to CM. The highest levels of IDPRs were in BRAF (45.95%), followed by PTEN (31.76%), NF1 (22.19%), c-KIT (21.82%), and NRAS (14.81%). Our STRING analysis found that each of these five proteins had more predicted interactions then expected (p-value < 1.0 × 10-16). Our analysis demonstrates that the mutations linked to CM likely affected IDPRs and possibly altered their highly complex PPIs. Quantifying IDPRs in BRAF, NRAS, c-KIT, NF1, and PTEN and understanding these protein regions are important processes as IDPRs can be possible drug targets for novel targeted therapies for treating CM.

Structure, domain organization, and different conformational states of stem cell factor-induced intact KIT dimers.

Using electron microscopy and fitting of crystal structures, we present the 3D reconstruction of ligand-induced dimers of intact receptor tyrosine kinase, KIT. We observe that KIT protomers form close contacts throughout the entire structure of ligand-bound receptor dimers, and that the dimeric receptors adopt multiple, defined conformational states. Interestingly, the homotypic interactions in the membrane proximal Ig-like domain of the extracellular region differ from those observed in the crystal structure of the unconstrained extracellular regions. We observe two prevalent conformations in which the tyrosine kinase domains interact asymmetrically. The asymmetric arrangement of the cytoplasmic regions may represent snapshots of molecular interactions occurring during trans autophosphorylation. Moreover, the asymmetric arrangements may facilitate specific intermolecular interactions necessary for trans phosphorylation of different KIT autophosphorylation sites that are required for stimulation of kinase activity and recruitment of signaling proteins by activated KIT.

Juxtaglomerular cell tumor: a morphological, immunohistochemical and genetic study of six cases.

Juxtaglomerular cell tumors (JGCTs) are rare tumors characterized by renin synthesis, hyperaldosteronism and hypertension. A curious immunohistochemical overlap between JGCT and gastrointestinal stromal tumor (GIST) including the expression of vimentin, CD34, CD117, α-smooth muscle actin was previously reported, prompting us to further investigate JGCT and its phenotypic and molecular genetic characteristics. Virtual karyotyping showed gain of chromosomes 3, 4, 10, 13, 17 and 18 in one JGCT, and fluorescence in situ hybridization (FISH) study confirmed this multiple gain pattern. Additionally, loss of chromosome 9 was observed in four of six cases analyzed with FISH. A whole genome expression analysis revealed 415 up-regulated (including renin, and CD117) and 325 down-regulated genes between the 2 cases. The study confirmed earlier reports on the gain of chromosomes 4 and 10, and provided further evidence of up-regulation of the genes located on these 2 chromosomes. For the first time our study indicated the importance of the loss of chromosome 9 and loss of expression of several tumor suppressor genes located on this chromosome as possible pathogenetic events important in development of JGCT.

Intestinal pacemaker C-KIT+ cells and synapses in allied Hirschsprung's disorders.

The cause of bowel dysmotility in allied Hirschsprung's disorders (AHDs) such as hypoganglionosis (HYPG), immature ganglia (IMG) and neuronal intestinal dysplasia (NID) remains unexplained. Recent experimental studies in mice have shown that c-kit gene product positive (C-KIT+) cells are responsible for intestinal pacemaker activity and that c-kit is also closely involved in synapse formation. To further understand the pathophysiology of AHDs, the authors used immunohistochemistry to study the distribution of C-KIT+ cells and synapses in the muscle layers of normal bowel from controls (12 cases) and bowel from patients with AHDs (10 patients; mean age, 3.0 years; 5 HYPG, 3 NID, 2 IMG). Anti-human C-KIT serum and monoclonal antibody 171B5 (a novel marker of synapses) were used for visualization of C-KIT+ cells and 171B5+ synapses, respectively. In normal bowel from controls and patients with AHDs, moderate to many C-KIT immunoreactive (C-KIT-IR+) cells were observed in the muscle layers. Myenteric plexuses were clearly demarcated by C-KIT-IR+ cells. 171B5 immunoreactive (171B5-IR+) synapses were abundant in the muscle layers and within the myenteric plexuses. In contrast, the number of C-KIT-IR+ cells or 171 B5-IR+ synapses was reduced in the muscle layers of bowel affected by AHDs, except within the myenteric plexuses, where there was a moderate to large number of 171B5-IR+ synapses identified. A lack of intestinal pacemaker C-KIT+ cells may be of great significance with respect to the bowel dysmotility associated with AHDs and also to the abnormal synapse formation seen in the muscle layers of bowel affected by these disorders.