PubMed Health⌕ Search

Biomedical subjects

R E Jensen

Publications and source records attributed to R E Jensen.

At least 19 recordsLinked to original sources
PubMed2001

UGO1 encodes an outer membrane protein required for mitochondrial fusion.

Membrane fusion plays an important role in controlling the shape, number, and distribution of mitochondria. In the yeast Saccharomyces cerevisiae, the outer membrane protein Fzo1p has been shown to mediate mitochondrial fusion. Using a novel genetic screen, we have isolated new mutants defective in the fusion of their mitochondria. One of these mutants, ugo1, shows several similarities to fzo1 mutants. ugo1 cells contain numerous mitochondrial fragments instead of the few long, tubular organelles seen in wild-type cells. ugo1 mutants lose mitochondrial DNA (mtDNA). In zygotes formed by mating two ugo1 cells, mitochondria do not fuse and mix their matrix contents. Fragmentation of mitochondria and loss of mtDNA in ugo1 mutants are rescued by disrupting DNM1, a gene required for mitochondrial division. We find that UGO1 encodes a 58-kD protein located in the mitochondrial outer membrane. Ugo1p appears to contain a single transmembrane segment, with its NH(2) terminus facing the cytosol and its COOH terminus in the intermembrane space. Our results suggest that Ugo1p is a new outer membrane component of the mitochondrial fusion machinery.

Amino Acid Motifs↗
PubMed2001

Mmm1p, a mitochondrial outer membrane protein, is connected to mitochondrial DNA (mtDNA) nucleoids and required for mtDNA stability.

In the yeast Saccharomyces cerevisiae, mitochondria form a branched, tubular reticulum in the periphery of the cell. Mmm1p is required to maintain normal mitochondrial shape and in mmm1 mutants mitochondria form large, spherical organelles. To further explore Mmm1p function, we examined the localization of a Mmm1p-green fluorescent protein (GFP) fusion in living cells. We found that Mmm1p-GFP is located in small, punctate structures on the mitochondrial outer membrane, adjacent to a subset of matrix-localized mitochondrial DNA nucleoids. We also found that the temperature-sensitive mmm1-1 mutant was defective in transmission of mitochondrial DNA to daughter cells immediately after the shift to restrictive temperature. Normal mitochondrial nucleoid structure also collapsed at the nonpermissive temperature with similar kinetics. Moreover, we found that mitochondrial inner membrane structure is dramatically disorganized in mmm1 disruption strains. We propose that Mmm1p is part of a connection between the mitochondrial outer and inner membranes, anchoring mitochondrial DNA nucleoids in the matrix.

Actins↗
PubMed2001

Division of mitochondria requires a novel DNM1-interacting protein, Net2p.

Mitochondria are dynamic organelles that undergo frequent division and fusion, but the molecular mechanisms of these two events are not well understood. Dnm1p, a mitochondria-associated, dynamin-related GTPase was previously shown to mediate mitochondrial fission. Recently, a genome-wide yeast two-hybrid screen identified an uncharacterized protein that interacts with Dnm1p. Cells disrupted in this new gene, which we call NET2, contain a single mitochondrion that consists of a network formed by interconnected tubules, similar to the phenotype of dnm1 Delta cells. NET2 encodes a mitochondria-associated protein with a predicted coiled-coil region and six WD-40 repeats. Immunofluorescence microscopy indicates that Net2p is located in distinct, dot-like structures along the mitochondrial surface, many of which colocalize with the Dnm1 protein. Fluorescence and immunoelectron microscopy shows that Dnm1p and Net2p preferentially colocalize at constriction sites along mitochondrial tubules. Our results suggest that Net2p is a new component of the mitochondrial division machinery.

Actins↗
PubMed2000

Yeast mitochondrial dynamics: fusion, division, segregation, and shape.

Mitochondria are essential organelles found in virtually all eukaryotic cells that play key roles in a variety of cellular processes. Mitochondria show a striking heterogeneity in their number, location, and shape in many different cell types. Although the dynamic nature of mitochondria has been known for decades, the molecules and mechanisms that mediate these processes are largely unknown. Recently, several laboratories have isolated and analyzed mutants in the yeast Saccharomyces cerevisiae defective in mitochondrial fusion and division, in the segregation of mitochondria to daughter cells, and in the establishment and maintenance of mitochondrial shape. These studies have identified several proteins that appear to mediate different aspects of mitochondrial morphogenesis. Although it is clear that many additional components have yet to be identified, some of the newly discovered proteins raise intriguing possibilities for how the processes of mitochondrial division, fusion, and segregation occur. Below we summarize our current understanding of the molecules known to be required for yeast mitochondrial dynamics.

Cell Division↗
PubMed2000

Two intermembrane space TIM complexes interact with different domains of Tim23p during its import into mitochondria.

Tim23p (translocase of the inner membrane) is an essential import component located in the mitochondrial inner membrane. To determine how the Tim23 protein itself is transported into mitochondria, we used chemical cross-linking to identify proteins adjacent to Tim23p during its biogenesis. In the absence of an inner membrane potential, Tim23p is translocated across the mitochondrial outer membrane, but not inserted into the inner membrane. At this intermediate stage, we find that Tim23p forms cross-linked products with two distinct protein complexes of the intermembrane space, Tim8p-Tim13p and Tim9p-Tim10p. Tim9p and Tim10p cross-link to the COOH-terminal domain of the Tim23 protein, which carries all of the targeting signals for Tim23p. Therefore, our results suggest that the Tim9p-Tim10p complex plays a key role in Tim23p import. In contrast, Tim8p and Tim13p cross-link to the hydrophilic NH(2)-terminal segment of Tim23p, which does not carry essential import information and, thus, the role of Tim8p-Tim13p is unclear. Tim23p contains two matrix-facing, positively charged loops that are essential for its insertion into the inner membrane. The positive charges are not required for interaction with the Tim9p-Tim10p complex, but are essential for cross-linking of Tim23p to components of the inner membrane insertion machinery, including Tim54p, Tim22p, and Tim12p.

Amino Acids↗
PubMed2000

Tim18p is a new component of the Tim54p-Tim22p translocon in the mitochondrial inner membrane.

The mitochondrial inner membrane contains two separate translocons: one required for the translocation of matrix-targeted proteins (the Tim23p-Tim17p complex) and one for the insertion of polytopic proteins into the mitochondrial inner membrane (the Tim54p-Tim22p complex). To identify new members of the Tim54p-Tim22p complex, we screened for high-copy suppressors of the temperature-sensitive tim54-1 mutant. We identified a new gene, TIM18, that encodes an integral protein of the inner membrane. The following genetic and biochemical observations suggest that the Tim18 protein is part of the Tim54p-Tim22p complex in the inner membrane: multiple copies of TIM18 suppress the tim54-1 growth defect; the tim18::HIS3 disruption is synthetically lethal with tim54-1; Tim54p and Tim22p can be coimmune precipitated with the Tim18 protein; and Tim18p, along with Tim54p and Tim22p, is detected in an approximately 300-kDa complex after blue native electrophoresis. We propose that Tim18p is a new component of the Tim54p-Tim22p machinery that facilitates insertion of polytopic proteins into the mitochondrial inner membrane.

Amino Acid Sequence↗
PubMed2000

Techniques and pitfalls of cervical lateral mass plate fixation.

We studied 35 successive patients who underwent subaxial lateral mass fixation between June 1994 and January 1996. Follow-up ranged from 2 to 3 1/2 years. Postoperative computed tomography scans showed 45 of 146 screws (31%) had suboptimal trajectory, but only 2 screws minimally impinged the foramen transversarium without adverse sequelae. The fusion rate was 89%. Study results show that optimal drilling technique and trajectory (15 degrees-25 degrees rostral in the sagittal plane, 20 degrees-30 degrees lateral in the axial plane) and intraoperative somatosensory evoked potential monitoring are associated with good screw placement and avoidance of complications, and bone grafting is associated with higher fusion rates.

Adult↗
PubMed1999

Division versus fusion: Dnm1p and Fzo1p antagonistically regulate mitochondrial shape.

In yeast, mitochondrial division and fusion are highly regulated during growth, mating and sporulation, yet the mechanisms controlling these activities are unknown. Using a novel screen, we isolated mutants in which mitochondria lose their normal structure, and instead form a large network of interconnected tubules. These mutants, which appear defective in mitochondrial division, all carried mutations in DNM1, a dynamin-related protein that localizes to mitochondria. We also isolated mutants containing numerous mitochondrial fragments. These mutants were defective in FZO1, a gene previously shown to be required for mitochondrial fusion. Surprisingly, we found that in dnm1 fzo1 double mutants, normal mitochondrial shape is restored. Induction of Dnm1p expression in dnm1 fzo1 cells caused rapid fragmentation of mitochondria. We propose that dnm1 mutants are defective in the mitochondrial division, an activity antagonistic to fusion. Our results thus suggest that mitochondrial shape is normally controlled by a balance between division and fusion which requires Dnm1p and Fzo1p, respectively.

Cell Division↗
PubMed1999

Protein translocation: is Hsp70 pulling my chain?

Hsp70 proteins in the lumen of the endoplasmic reticulum and in the mitochondrial matrix are thought to drive the translocation of proteins into each organelle. Recent experiments aimed at distinguishing between two models for Hsp70 function appear to reach opposite conclusions.

Animals↗
PubMed1998

Tim23p contains separate and distinct signals for targeting to mitochondria and insertion into the inner membrane.

The Tim23 protein is an essential inner membrane (IM) component of the yeast mitochondrial protein import pathway. Tim23p does not carry an amino-terminal presequence; therefore, the targeting information resides within the mature protein. Tim23p is anchored in the IM via four transmembrane segments and has two positively charged loops facing the matrix. To identify the import signal for Tim23p, we have constructed several altered versions of the Tim23 protein and examined their function and import in yeast cells, as well as their import into isolated mitochondria. We replaced the positively charged amino acids in one or both loops with alanine residues and found that the positive charges are not required for import into mitochondria, but at least one positively charged loop is required for insertion into the IM. Furthermore, we find that the signal to target Tim23p to mitochondria is carried in at least two of the hydrophobic transmembrane segments. Our results suggest that Tim23p contains separate import signals: hydrophobic segments for targeting Tim23p to mitochondria, and positively charged loops for insertion into the IM. We therefore propose that Tim23p is imported into mitochondria in at least two distinct steps.

Binding Sites↗
PubMed1998

Characterization of the mitochondrial inner membrane translocase complex: the Tim23p hydrophobic domain interacts with Tim17p but not with other Tim23p molecules.

Tim23p is a mitochondrial inner membrane protein essential for the import of proteins from the cytosol. Tim23p contains an amino-terminal hydrophilic segment and a carboxyl-terminal hydrophobic domain (Tim23Cp). To study the functions and interactions of the two parts of Tim23p separately, we constructed tim23N, encoding only the hydrophilic region of Tim23p, and tim23C, encoding only the hydrophobic domain of Tim23p. Only the Tim23C protein is imported into mitochondria, indicating that the mitochondrial targeting information in Tim23p resides in its membrane spans or intervening loops. Tim23Cp, however, cannot substitute for full-length Tim23p, suggesting that the hydrophilic portion of Tim23p also performs an essential function in mitochondrial protein import. We found that overexpression of Tim23Cp is toxic to yeast cells that carry the tim23-1 mutation. Excess Tim23Cp causes Tim23-1p to disappear, leaving tim23-1 cells without a full-length version of the Tim23 protein. If Tim17p, another inner membrane import component, is overexpressed along with Tim23Cp, the toxicity of Tim23Cp is largely reversed and the Tim23-1 protein no longer disappears. In coimmunoprecipitations from solubilized mitochondria, Tim17p associates with the Tim23C protein. In addition, we show that Tim23p and Tim17p can be chemically cross-linked to each other in intact mitochondria. We conclude that the hydrophobic domain encoded by tim23C targets Tim23p to the mitochondria and mediates the direct interaction between Tim23p and Tim17p. In contrast, Tim23Cp cannot be coimmunoprecipitated with Tim23p, raising the possibility that the hydrophobic domain of Tim23p does not interact with other Tim23 molecules.

Carrier Proteins↗
PubMed1998

The effectiveness of gallium citrate Ga 67 radionuclide imaging in vertebral osteomyelitis revisited.

We investigated the role of gallium citrate Ga 67 scanning in diagnosing spondylodiscitis. Scans of 41 patients with suspected spondylodiscitis showed increased radionuclide uptake in 39 patients; these findings correlated with those of magnetic resonance imaging and were proved by biopsy. Two patients with negative findings on gallium scans had been strongly suspected of having spondylodiscitis; biopsy findings in these patients showed degenerative changes. Thirteen patients had negative cultures, while 22 had polygenic infections and 4 had granulomatous infections. Gallium scanning proved to be 100% sensitive, specific, and accurate. The interrater accuracy was excellent. Follow-up scans were used to track therapeutic progress. We recommend complementary bone and gallium scans in cases of suspected spinal infections. If the scan is positive, a biopsy should be done. If the scans are negative, no further investigation is needed.

Adolescent↗
PubMed1998

Percutaneous transpedicular discectomy and drainage in pyogenic spondylodiscitis.

Spondylodiscitis normally heals itself, but it can cause bone destruction leading to deformity and often pain. Debridement of these infections by percutaneous transpedicular discectomy with access from adjacent caudal pedicles can accelerate natural healing and prevent progression to bone destruction and deformity. We outline this technique and discuss a series of 28 patients treated using a percutaneous transpedicular approach to obtain culture and histopathology specimens, permit drainage and antibiotic irrigation, and provide a channel for granulation tissue to invade the infected space. This procedure is safe and effective, but it is contraindicated for epidural abscess or granulation tissue-induced neurocompression and it is ineffective against extensive bone destruction.

Adolescent↗
PubMed1997

The Tim54p-Tim22p complex mediates insertion of proteins into the mitochondrial inner membrane.

We have identified a new protein, Tim54p, located in the yeast mitochondrial inner membrane. Tim54p is an essential import component, required for the insertion of at least two polytopic proteins into the inner membrane, but not for the translocation of precursors into the matrix. Several observations suggest that Tim54p and Tim22p are part of a protein complex in the inner membrane distinct from the previously characterized Tim23p-Tim17p complex. First, multiple copies of the TIM22 gene, but not TIM23 or TIM17, suppress the growth defect of a tim54-1 temperature-sensitive mutant. Second, Tim22p can be coprecipitated with Tim54p from detergent-solubilized mitochondria, but Tim54p and Tim22p do not interact with either Tim23p or Tim17p. Finally, the tim54-1 mutation destabilizes the Tim22 protein, but not Tim23p or Tim17p. Our results support the idea that the mitochondrial inner membrane carries two independent import complexes: one required for the translocation of proteins across the inner membrane (Tim23p-Tim17p), and the other required for the insertion of proteins into the inner membrane (Tim54p-Tim22p).

Biological Transport↗
PubMed1997

Tim23, a protein import component of the mitochondrial inner membrane, is required for normal activity of the multiple conductance channel, MCC.

We previously showed that the conductance of a mitochondrial inner membrane channel, called MCC, was specifically blocked by peptides corresponding to mitochondrial import signals. To determine if MCC plays a role in protein import, we examined the relationship between MCC and Tim23p, a component of the protein import complex of the mitochondrial inner membrane. We find that antibodies against Tim23p, previously shown to inhibit mitochondrial protein import, inhibit MCC activity. We also find that MCC activity is altered in mitochondria isolated from yeast carrying the tim23-1 mutation. In contrast to wild-type MCC, we find that the conductance of MCC from the tim23-1 mutant is not significantly blocked by mitochondrial presequence peptides. Tim23 antibodies and the tim23-1 mutation do not, however, alter the activity of PSC, a presequence-peptide sensitive channel in the mitochondrial outer membrane. Our results show that Tim23p is required for normal MCC activity and raise the possibility that precursors are translocated across the inner membrane through the pore of MCC.

Amino Acid Sequence↗
PubMed1997

The mitochondrial protein import pathway: are precursors imported through membrane channels?

Mitochondrial biogenesis requires the import of hundreds of different proteins from the cytosol. Protein import into mitochondria is a multistep pathway that includes recognition of precursor proteins by machinery both in the cytoplasm and on the mitochondrial surface, translocation of the precursor across one or both mitochondrial membranes, and folding of the protein after its import into the organelle. Over the past several years, many components of the import machinery have been identified using both biochemical and genetic methods. Recently, significant progress has been made determining the function of some of these import proteins. One purpose of this minireview is to summarize our current understanding of the import pathway, and to introduce the topics of the minireviews that will follow. The other goal of this minireview is to discuss recent findings suggesting that proteins are translocated across both the mitochondrial inner and outer membranes through aqueous channels.

Cell Membrane↗