Tentorial Venous Anatomy: Variation in the Healthy Population.

BACKGROUND AND PURPOSE: A new transtentorial venous system consisting of medial, intermediate, and lateral tentorial veins, connecting infra- and supratentorial compartments, was recently shown in 2 cadaver dissections and 2 patient scans. We sought to characterize the venous patterns within the tentorium and their relation to measures of skull development in a cohort of healthy adults. MATERIALS AND METHODS: We retrospectively reviewed tentorial venous anatomy of the head using CTA/CTV performed for routine care or research purposes in 238 patients. Included studies had adequate contrast opacification of venous structures and a section thickness of ≤2 mm; we excluded cases with space-occupying lesions and vascular pathologies. Tentorial angle, dural sinus configurations, and measures of skull base development were assessed as predictors of tentorial venous anatomy variation via Cramér V association, the binary encoded Pearson correlation, and nearest-point algorithm with the Euclidean distance metric for clustering. RESULTS: Tentorial vein development was related to the ringed configuration of the tentorial sinuses (P < .005). There were 3 configurations. Groups 1A and 1B (n = 50/238) had ringed configuration, while group 2 did not (n = 188/238). Group 1A (n = 38/50) had a medialized ringed configuration, and group 1B had a lateralized ringed configuration (n = 12/50). Measurements of skull base development were predictive of these groups. The ringed configuration of group 1 was related to the presence of a split confluens, which correlated with a decreased internal auditory canal-petroclival fissure angle. Configuration 1A was related to the degree of petrous apex pneumatization (P value = .010). CONCLUSIONS: Variations in the transtentorial venous system directly correlate with cranial development.

Autoproteolysis in nucleoporin biogenesis.

We have molecularly characterized a proteolytic cleavage in conserved nuclear pore complex proteins. This cleavage, previously demonstrated to be essential for the biogenesis of two nuclear pore complex proteins in mammals (Nup98 and Nup96) and yeast (Nup145-N and Nup145-C), occurs between Phe and Ser residues within a highly conserved domain in a polyprotein precursor. Here, we show that a protease is not involved in the cleavage event. By using a combination of domain mapping and site-directed mutagenesis, we demonstrate that the human nuclear pore complex protein Nup98 specifically cleaves itself between F863 and S864. A region of Nup98, amino acids 715-920, is able to cleave, whereas a smaller region, amino acids 772-920, does not cleave. In addition, we have generated a Nup98 mutant that cleaves under defined conditions in vitro. Further, the two cleaved fragments of Nup98 form a complex, providing a possible mechanism whereby specific, yet low-affinity, binding between Nup98 and Nup96 is responsible for the nuclear targeting of Nup96. Although apparently unrelated evolutionarily, Nup98 has converged on an autoproteolytic biogenesis mechanism similar to that of hedgehog proteins, the inteins, and the N-terminal nucleophile proteins.

Nuclear import of the TATA-binding protein: mediation by the karyopherin Kap114p and a possible mechanism for intranuclear targeting.

Binding of the TATA-binding protein (TBP) to the promoter is the first and rate limiting step in the formation of transcriptional complexes. We show here that nuclear import of TBP is mediated by a new karyopherin (Kap) (importin) family member, Kap114p. Kap114p is localized to the cytoplasm and nucleus. A complex of Kap114p and TBP was detected in the cytosol and could be reconstituted using recombinant proteins, suggesting that the interaction was direct. Deletion of the KAP114 gene led to specific mislocalization of TBP to the cytoplasm. We also describe two other potential minor import pathways for TBP. Consistent with other Kaps, the dissociation of TBP from Kap114p is dependent on RanGTP. However, we could show that double stranded, TATA-containing DNA stimulates this RanGTP-mediated dissociation of TBP, and is necessary at lower RanGTP concentrations. This suggests a mechanism where, once in the nucleus, TBP is preferentially released from Kap114p at the promoter of genes to be transcribed. In this fashion Kap114p may play a role in the intranuclear targeting of TBP.

A novel nuclear import pathway for the transcription factor TFIIS.

We have identified a novel pathway for protein import into the nucleus. We have shown that the previously identified but uncharacterized yeast protein Nmd5p functions as a karyopherin. It was therefore designated Kap119p (karyopherin with Mr of 119 kD). We localized Kap119p to both the nucleus and the cytoplasm. We identified the transcription elongation factor TFIIS as its major cognate import substrate. The cytoplasmic Kap119p exists as an approximately stoichiometric complex with TFIIS. RanGTP, not RanGDP, dissociated the isolated Kap119p/TFIIS complex and bound to Kap119p. Kap119p also bound directly to a number of peptide repeat containing nucleoporins in overlay assays. In wild-type cells, TFIIS was primarily localized to the nucleus. In a strain where KAP119 has been deleted, TFIIS was mislocalized to the cytoplasm indicating that TFIIS is imported into the nucleus by Kap119p. The transport of various substrates that use other karyopherin-mediated import or export pathways was not affected in a kap119Delta strain. Hence Kap119p is a novel karyopherin that is responsible for the import of the transcription elongation factor TFIIS.

Nuclear import and the evolution of a multifunctional RNA-binding protein.

La (SS-B) is a highly expressed protein that is able to bind 3'-oligouridylate and other common RNA sequence/structural motifs. By virtue of these interactions, La is present in a myriad of nuclear and cytoplasmic ribonucleoprotein complexes in vivo where it may function as an RNA-folding protein or RNA chaperone. We have recently characterized the nuclear import pathway of the S. cerevisiae La, Lhp1p. The soluble transport factor, or karyopherin, that mediates the import of Lhp1p is Kap108p/Sxm1p. We have now determined a 113-amino acid domain of Lhp1p that is brought to the nucleus by Kap108p. Unexpectedly, this domain does not coincide with the previously identified nuclear localization signal of human La. Furthermore, when expressed in Saccharomyces cerevisiae, the nuclear localization of Schizosaccharomyces pombe, Drosophila, and human La proteins are independent of Kap108p. We have been able to reconstitute the nuclear import of human La into permeabilized HeLa cells using the recombinant human factors karyopherin alpha2, karyopherin beta1, Ran, and p10. As such, the yeast and human La proteins are imported using different sequence motifs and dissimilar karyopherins. Our results are consistent with an intermingling of the nuclear import and evolution of La.

Transport routes through the nuclear pore complex.

The nuclear pore complex can be considered to be the stationary phase of bidirectional traffic between the nucleus and the cytoplasm. The mobile phase consists of karyopherins, transport substrates, and the small GTPase Ran and its modulators. Recently, the family of karyopherins was expanded with the recognition of numerous open reading frames with limited homology to karyopherin beta 1. In several cases, the specific substrates transported by the new karyopherins have been identified, allowing the characterization of new pathways into and out of the nucleus. However, the mechanisms of transport, particularly the role of Ran, remain poorly understood.

A distinct and parallel pathway for the nuclear import of an mRNA-binding protein.

Three independent pathways of nuclear import have so far been identified in yeast, each mediated by cognate nuclear transport factors, or karyopherins. Here we have characterized a new pathway to the nucleus, mediated by Mtr10p, a protein first identified in a screen for strains defective in polyadenylated RNA export. Mtr10p is shown to be responsible for the nuclear import of the shuttling mRNA-binding protein Npl3p. A complex of Mtr10p and Npl3p was detected in cytosol, and deletion of Mtr10p was shown to lead to the mislocalization of nuclear Npl3p to the cytoplasm, correlating with a block in import. Mtr10p bound peptide repeat-containing nucleoporins and Ran, suggesting that this import pathway involves a docking step at the nuclear pore complex and is Ran dependent. This pathway of Npl3p import is distinct and does not appear to overlap with another known import pathway for an mRNA-binding protein. Thus, at least two parallel pathways function in the import of mRNA-binding proteins, suggesting the need for the coordination of these pathways.

A nuclear import pathway for a protein involved in tRNA maturation.

A limited number of transport factors, or karyopherins, ferry particular substrates between the cytoplasm and nucleoplasm. We identified the Saccharomyces cerevisiae gene YDR395w/SXM1 as a potential karyopherin on the basis of limited sequence similarity to known karyopherins. From yeast cytosol, we isolated Sxm1p in complex with several potential import substrates. These substrates included Lhp1p, the yeast homologue of the human autoantigen La that has recently been shown to facilitate maturation of pre-tRNA, and three distinct ribosomal proteins, Rpl16p, Rpl25p, and Rpl34p. Further, we demonstrate that Lhp1p is specifically imported by Sxm1p. In the absence of Sxm1p, Lhp1p was mislocalized to the cytoplasm. Sxm1p and Lhp1p represent the karyopherin and a cognate substrate of a unique nuclear import pathway, one that operates upstream of a major pathway of pre-tRNA maturation, which itself is upstream of tRNA export in wild-type cells. In addition, through its association with ribosomal proteins, Sxm1p may have a role in coordinating ribosome biogenesis with tRNA processing.

On signal sequence polymorphisms and diseases of distribution.

We report a previously unappreciated property of the signals that target organelle-specific proteins to their subcellular sites of action. Such targeting sequences are shown to be polymorphic. We discovered this polymorphism when we cloned the mitochondrial manganese-containing superoxide dismutase from cell lines of normal individuals and patients with genetic diseases of premature aging and compared their sequences to each other and to those previously reported. The polymorphism consists of a single nucleotide change in the region of the DNA that encodes the signal sequence such that either an alanine or valine is present. Subsequently, eight cell lines were analyzed and all three possible combinations of the two signal sequences were observed. Such signal sequence polymorphisms could result in diseases of distribution, where essential proteins are not properly targeted, thereby leading to absolute or relative deficiencies of critical enzymes within specific cellular compartments. Progeria and related syndromes may be diseases of distribution.

Direct selection of antibodies that coordinate metals from semisynthetic combinatorial libraries.

An iterative strategy for the selection of catalytic metalloantibodies is described. The first stage of this strategy is validated by the selection of semisynthetic antibodies that coordinate a variety of different metal ions and the metal oxide magnetite. These results have implications not only for the development of catalytic metalloantibodies but also for the development of reagents for magnetic resonance imaging, delivery of radioisotopes, and purification of recombinant proteins.

Oxygen distribution in squamous cell carcinoma metastases and its relationship to outcome of radiation therapy.

Oxygen distribution was measured in 31 fixed lymph node metastases (mean diameter 4.4 cm +/- 0.8 cm) from squamous cell carcinoma of the head and neck by passing a needle electrode through each tumor under CT guidance. Thirteen tumors had uniform oxygen distribution with all measurements under 10 mm Hg. Six tumors had uniform oxygen distribution with all measurements above 10 mm Hg, and twelve tumors had variable oxygen distribution with measurements higher in the periphery than in the center. Response to radiation therapy was judged by changes in tumor volume 90 days following completion of therapy compared to pre-therapy volume. Eighteen tumors were considered complete responders (CR); eleven, non-responders (NR); two, partial responders (PR). No statistically significant difference in radiation dose or tumor size was seen in the PR and CR groups. Mean pO2 was 20.6 (+/- 4.4) mm Hg in the CR group and 4.7 (+/- 3.0) mm Hg in the NR group (p less than 0.001). Normalized pO2 content defined as the total tumor oxygen content normalized by dividing by the volume was 37.4 (+/- 8.2) mm Hg in the CR group and 8.2 (+/- 5.1) mm Hg in the NR group (p less than 0.001). The volume and oxygen levels of each tumor were tabulated and analyzed. Twelve tumors had greater than 26% of their volume containing a pO2 less than 8 mm Hg. Eleven of these were NR and one PR. Nineteen tumors had less than 26% of their volume containing a pO2 less than 8 mm Hg. Eighteen were CR and one PR (p less than 0.001). These data suggest that oxygen plays a significant role in human tumor response to radiation therapy. Oxygen measurements appear to allow separation of subgroups of patients with a poor prognosis who would most benefit from maneuvers to circumvent the hypoxic effect.

CT-guided laser therapy in resistant human tumors: phase I clinical trials.

Photodynamic therapy was performed on ten tumors in patients who did not respond to initial therapy and for whom no additional conventional therapy was available. A sensitizing agent (hematoporphyrin derivative [HpD]) was injected directly into each tumor under computed tomographic (CT) guidance to deliver high concentrations to the tumor and to minimize systemic toxicity. Three to 6 days after the injection, a clear Teflon sheath catheter was placed into the tumor under CT guidance. The tumor was exposed to red light (630-nm wavelength) through laser fiberoptics inserted in the sheath. The initial investigation confirmed the technical feasibility of CT-guided photodynamic therapy by means of intratumoral HpD injections and laser exposure through fiberoptics inserted in sheath catheters. The toxicity from a single treatment was minor, and the tumor response was encouraging.

Esophageal speech: double-contrast evaluation of the pharyngo-esophageal segment.

Alaryngeal voice is usually accompanied by esophageal speech; however, about 40% of laryngectomy patients are unable to achieve adequate esophageal speech and must rely on mechanical devices for communication. A technique was developed for performing double-contrast studies of the hypopharynx, cervical esophagus, and pharyngo-esophageal segment using thick barium and the air normally injected for speech. Simultaneous audio and video recordings obtained during esophageal speech allowed correlation of the quality of speech with the motion of the pharyngo-esophageal segment. In 35 patients with various degrees of fluency in esophageal speech, normal and abnormal function of the pharyngo-esophageal segment was documented. Inadequate esophageal speech can be related to abnormal motion of the pharyngo-esophageal segment.

Cryptosporidiosis of the stomach and small intestine in patients with AIDS.

Cryptosporidiosis is a parasitic disease of the gastrointestinal tract that causes a choleralike diarrhea in patients with the acquired immunodeficiency syndrome (AIDS). Review of barium studies of the stomach and small intestine in 16 AIDS patients with cryptosporidiosis showed that the studies were abnormal in 13. Of these, five had moderate or marked prominence of the mucosal folds, and eight had slight prominence. The jejunum was predominantly involved in nine; the entire small intestine was uniformly affected in the other four. Three of the 16 patients had moderate or marked dilatation of the small intestine. One had marked dilution of the barium, and one had marked fragmentation and flocculation of the barium. There was narrowing and rigidity of the gastric antrum in two patients. These radiographic findings are nonspecific, but are indicative of cryptosporidiosis in a patient with AIDS and protracted diarrhea.