Cooperation between p53 and the telomere-protecting shelterin component Pot1a in endometrial carcinogenesis.

Type II endometrial cancer (EMCA) represents only 10% of all EMCAs, but accounts for 40% of EMCA-related mortality. Previous studies of human tumors have shown an association between Type II tumors and damaged telomeres. We hypothesized that the lack of murine Type II EMCA models is due to the extremely long telomeres in laboratory mouse strains. We previously showed that telomerase-null mice with critically short telomeres developed endometrial lesions histologically resembling endometrial intraepithelial carcinoma (EIC), the accepted precursor for Type II EMCA. However, these mice did not develop invasive endometrial adenocarcinoma, and instead succumbed prematurely to multi-organ failure. Here, we modeled critical telomere attrition by conditionally inactivating Pot1a, a component of the shelterin complex that stabilizes telomeres, within endometrial epithelium. Inactivation of Pot1a by itself did not stimulate endometrial carcinogenesis, and did not result in detectable DNA damage or apoptosis in endometrium. However, simultaneous inactivation of Pot1a and p53 resulted in EIC-like lesions by 9 months indistinguishable from those seen in late generation telomerase-null mice. These lesions progressed to invasive endometrial adenocarcinomas as early as 9 months of age with metastatic disease in 100% of the animals by 15 months. These tumors were poorly differentiated endometrial adenocarcinomas with prominent nuclear atypia, resembling human Type II cancers. Furthermore, these tumors were aneuploid with double-stranded DNA breaks and end-to-end telomere fusions and most were tetraploid or near-tetraploid. These studies lend further support to the hypothesis that telomeric instability has a critical role in Type II endometrial carcinogenesis and provides an intriguing in-vivo correlate to recent studies implicating telomere-dependent tetraploidization as an important mechanism in carcinogenesis.

What determines growth direction in fungal hyphae?

We used high-resolution video microscopy and image analysis to map the trajectory of the Spitzenkörper in growing hyphae of Neurospora crassa and to correlate it with growth directionality. The Spitzenkörper followed a tortuous trajectory produced by a dominant forward motion accompanied by frequent, transverse oscillations. In hyphae with a fixed growth direction, the regression line of the Spitzenkörper trajectory corresponded to the longitudinal axis of the hypha. A permanent change in growth direction, i.e., the establishment of a new growth axis, was correlated with a sustained shift in Spitzenkörper trajectory away from the existing cell axis. In meandering hyphae, changes in growth directionality occurred somewhat erratically but there was a strong compensatory tendency reversing directional shifts and maintaining an overall fixed direction of growth. Although external factors greatly affect hyphal growth direction (tropisms), they are probably not the primary determinants of growth directionality. Inhibitors of microtubules, but not of actin microfilaments, caused hyphae to lose their growth directionality-providing support for the idea that Spitzenkörper trajectory is determined internally by a growing scaffolding of cytoplasmic microtubules. The meandering morphology of N. crassa hyphae was duplicated by computer simulation in support of the idea that hyphal morphogenesis is controlled by the position of the Spitzenkörper functioning as a vesicle supply center.

Analysis of the role of the Spitzenkörper in fungal morphogenesis by computer simulation of apical branching in Aspergillus niger.

High-resolution video microscopy, image analysis, and computer simulation were used to study the role of the Spitzenkörper (Spk) in apical branching of ramosa-1, a temperature-sensitive mutant of Aspergillus niger. A shift to the restrictive temperature led to a cytoplasmic contraction that destabilized the Spk, causing its disappearance. After a short transition period, new Spk appeared where the two incipient apical branches emerged. Changes in cell shape, growth rate, and Spk position were recorded and transferred to the FUNGUS SIMULATOR program to test the hypothesis that the Spk functions as a vesicle supply center (VSC). The simulation faithfully duplicated the elongation of the main hypha and the two apical branches. Elongating hyphae exhibited the growth pattern described by the hyphoid equation. During the transition phase, when no Spk was visible, the growth pattern was nonhyphoid, with consecutive periods of isometric and asymmetric expansion; the apex became enlarged and blunt before the apical branches emerged. Video microscopy images suggested that the branch Spk were formed anew by gradual condensation of vesicle clouds. Simulation exercises where the VSC was split into two new VSCs failed to produce realistic shapes, thus supporting the notion that the branch Spk did not originate by division of the original Spk. The best computer simulation of apical branching morphogenesis included simulations of the ontogeny of branch Spk via condensation of vesicle clouds. This study supports the hypothesis that the Spk plays a major role in hyphal morphogenesis by operating as a VSC-i.e., by regulating the traffic of wall-building vesicles in the manner predicted by the hyphoid model.

Apical branching in a temperature sensitive mutant of Aspergillus niger.

An apical branching, temperature-sensitive, mutant of Aspergillus niger (ramosa-1) was isolated by UV mutagenesis. Ramosa-1 has a wild type morphology at 23 degrees C, but branches apically when shifted to 34 degrees C. The cytological events leading to apical branching were recorded by video-enhanced phase contrast microscopy. The first event was a momentary, localized, cytoplasmic contraction lasting approximately 1 s. This contraction was seen as a sudden unidirectional movement of visible organelles (mitochondria, spheroid bodies) toward the hyphal apex. During the contraction, there was a transitory sharp increase in refractive index in a localized area of cytoplasm in the apex or subapex of the cell. Within 5 s, the Spitzenkörper retracted from its normal position next to the apical pole and disappeared from view 20 to 50 s later. Hyphal elongation rate diminished sharply, and the typical distribution of organelles at the hyphal tip was disturbed. After 210-240 s, organelle distribution returned to normal, polarized growth resumed, but instead of one Spitzenkörper two new Spitzenkörper appeared, each giving rise to an apical branch. The second branch Spitzenkörper appeared with a 60- to 100-s delay. We did not observe the original Spitzenkörper dividing in two; instead, the new Spitzenkörper arose de novo from vesicle clouds that formed in the apical region next to the future site of branch emergence. In all instances that we examined, the dislocation and disappearance of the Spitzenkörper was preceded by cytoplasmic contractions. We therefore suspect the existence of an intimate connection between the cytoskeletal network and the Spitzenkörper. Accordingly, we propose that the apical branching phenotype in ramosa-1 is triggered by a molecular event that induces a transient alteration in cytoskeleton organization.

[Renal malacoplakia in childhood. Report of a case]. / Malacoplaquia renal en la infancia. Informe de un caso.

Renal malacoplakia is a rare entity with less frequency in children. In the past 7 years it was the first case reported in a third level regional hospital. The present clinical case showed chronic urinary tract infection and IVP showed right renal exclusion. He was nephrectomized on the right side because of arterial hypertension which started in the third year of the illness. The diagnosis was made after the surgery by microscopic examination, with the characteristic Michaelis-Gutmann bodies being shown.