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Y Lao

Publications and source records attributed to Y Lao.

11 recordsLinked to original sources
PubMed2001

The C2B domain of synaptotagmin I is a Ca2+-binding module.

Synaptotagmin I is a synaptic vesicle protein that contains two C(2) domains and acts as a Ca(2+) sensor in neurotransmitter release. The Ca(2+)-binding properties of the synaptotagmin I C(2)A domain have been well characterized, but those of the C(2)B domain are unclear. The C(2)B domain was previously found to pull down synaptotagmin I from brain homogenates in a Ca(2+)-dependent manner, leading to an attractive model whereby Ca(2+)-dependent multimerization of synaptotagmin I via the C(2)B domain participates in fusion pore formation. However, contradictory results have been described in studies of Ca(2+)-dependent C(2)B domain dimerization, as well as in analyses of other C(2)B domain interactions. To shed light on these issues, the C(2)B domain has now been studied using biophysical techniques. The recombinant C(2)B domain expressed as a GST fusion protein and isolated by affinity chromatography contains tightly bound bacterial contaminants despite being electrophoretically pure. The contaminants bind to a polybasic sequence that has been previously implicated in several C(2)B domain interactions, including Ca(2+)-dependent dimerization. NMR experiments show that the pure recombinant C(2)B domain binds Ca(2+) directly but does not dimerize upon Ca(2+) binding. In contrast, a cytoplasmic fragment of native synaptotagmin I from brain homogenates, which includes the C(2)A and C(2)B domains, participates in a high molecular weight complex as a function of Ca(2+). These results show that the recombinant C(2)B domain of synaptotagmin I is a monomeric, autonomously folded Ca(2+)-binding module and suggest that a potential function of synaptotagmin I multimerization in fusion pore formation does not involve a direct interaction between C(2)B domains or requires a posttranslational modification.

Amino Acid Motifs↗
PubMed2001

Synaptotagmin VII as a plasma membrane Ca(2+) sensor in exocytosis.

Synaptotagmins I and II are Ca(2+) binding proteins of synaptic vesicles essential for fast Ca(2+)-triggered neurotransmitter release. However, central synapses and neuroendocrine cells lacking these synaptotagmins still exhibit Ca(2+)-evoked exocytosis. We now propose that synaptotagmin VII functions as a plasma membrane Ca(2+) sensor in synaptic exocytosis complementary to vesicular synaptotagmins. We show that alternatively spliced forms of synaptotagmin VII are expressed in a developmentally regulated pattern in brain and are concentrated in presynaptic active zones of central synapses. In neuroendocrine PC12 cells, the C(2)A and C(2)B domains of synaptotagmin VII are potent inhibitors of Ca(2+)-dependent exocytosis, but only when they bind Ca(2+). Our data suggest that in synaptic vesicle exocytosis, distinct synaptotagmins function as independent Ca(2+) sensors on the two fusion partners, the plasma membrane (synaptotagmin VII) versus synaptic vesicles (synaptotagmins I and II).

Aging↗
PubMed2000

Replication protein A interactions with DNA. III. Molecular basis of recognition of damaged DNA.

Human replication protein A (RPA) is a heterotrimeric single-stranded DNA-binding protein (subunits of 70, 32, and 14 kDa) that is required for cellular DNA metabolism. RPA has been reported to interact specifically with damaged double-stranded DNA and to participate in multiple steps of nucleotide excision repair (NER) including the damage recognition step. We have examined the mechanism of RPA binding to both single-stranded and double-stranded DNA (ssDNA and dsDNA, respectively) containing damage. We show that the affinity of RPA for damaged dsDNA correlated with disruption of the double helix by the damaged bases and required RPAs ssDNA-binding activity. We conclude that RPA is recognizing single-stranded character caused by the damaged nucleotides. We also show that RPA binds specifically to damaged ssDNA. The specificity of binding varies with the type of damage with RPA having up to a 60-fold preference for a pyrimidine(6-4)pyrimidone photoproduct. We show that this specific binding was absolutely dependent on the zinc-finger domain in the C-terminus of the 70-kDa subunit. The affinity of RPA for damaged ssDNA was 5 orders of magnitude higher than that of the damage recognition protein XPA (xeroderma pigmentosum group A protein). These findings suggest that RPA probably binds to both damaged and undamaged strands in the NER excision complex. RPA binding may be important for efficient excision of damaged DNA in NER.

Conserved Sequence↗
PubMed2000

Bilateral papilloedema associated with lumbo-sacral intraspinal tumor.

OBJECTIVE: To study the possible mechanism of bilateral papilloedema associated with intraspinal tumor at the lumbo-sacral level. METHODS: Three cases were reported. Detailed clinical history was evaluated. Patients were followed up for 2, 3 and 5 months after resection of the tumor. Related literature was reviewed. RESULTS: Papilloedema was soon resolved and the level of protein in the cerebrospinal fluid (CSF) decreased significantly after resection of tumor. These indicated a close relationship between papilloedema, CSF protein and intraspinal tumor. CONCLUSION: This is the first report of papilloedema associated with a lumbo-sacral intraspinal tumor in Chinese literature. The high level of protein in CSF may be the cause of both papilloedema and increased intracranial pressure. In patients with bilateral papilloedema, if no intracranial cause could be found, lumbo-sacral intraspinal tumor should be considered.

Adult↗
PubMed2000

[Poly-static quantitative perimetry for detection of open angle glaucoma].

OBJECTIVE: To evaluate the sensitivity of poly-static quantitative perimetry for the detection of visual field defect in open angle glaucoma. METHODS: 48 patients (95 eyes) of open angle glaucoma were examined with poly-static quantitative perimetry [Friedmann visual field analyzer (FVFA)] and dynamic perimetry (tangent screen) respectively. RESULTS: 26 eyes showed small defect of visual field on tangent screen with 2/1,000 visual size, and 23 eyes (88%) showed the same defects checked with FVFA; visual field defects of early stage glaucoma were found in 26 eyes checked with FVFA, but only 16 eyes showed the same defects on tangent screen with 1/1,000 visual size; the visual field defects of glaucoma on tangent screen in 43 eyes were found larger when they were checked with FVFA. CONCLUSION: Comparing with dynamic perimetry, poly-static quantitative perimetry is more sensitive to detect visual field defect of early stage glaucoma.

Adult↗
PubMed1999

Replication protein A interactions with DNA. 1. Functions of the DNA-binding and zinc-finger domains of the 70-kDa subunit.

Human replication protein A (RPA) is a multiple subunit single-stranded DNA-binding protein that is required for multiple processes in cellular DNA metabolism. This complex, composed of subunits of 70, 32, and 14 kDa, binds to single-stranded DNA (ssDNA) with high affinity and participates in multiple protein-protein interactions. The 70-kDa subunit of RPA is known to be composed of multiple domains: an N-terminal domain that participates in protein interactions, a central DNA-binding domain (composed of two copies of a ssDNA-binding motif), a putative (C-X2-C-X13-C-X2-C) zinc finger, and a C-terminal intersubunit interaction domain. A series of mutant forms of RPA were used to elucidate the roles of these domains in RPA function. The central DNA-binding domain was necessary and sufficient for interactions with ssDNA; however, adjacent sequences, including the zinc-finger domain and part of the N-terminal domain, were needed for optimal ssDNA-binding activity. The role of aromatic residues in RPA-DNA interactions was examined. Mutation of any one of the four aromatic residues shown to interact with ssDNA had minimal effects on RPA activity, indicating that individually these residues are not critical for RPA activity. Mutation of the zinc-finger domain altered the structure of the RPA complex, reduced ssDNA-binding activity, and eliminated activity in DNA replication.

Antigens, Viral, Tumor↗
PubMed1999

Replication protein A interactions with DNA. 2. Characterization of double-stranded DNA-binding/helix-destabilization activities and the role of the zinc-finger domain in DNA interactions.

Human replication protein A (RPA) is a heterotrimeric single-stranded DNA-binding protein that is composed of subunits of 70, 32, and 14 kDa. RPA is required for multiple processes in cellular DNA metabolism. RPA has been reported to (1) bind with high affinity to single-stranded DNA (ssDNA), (2) bind specifically to certain double-stranded DNA (dsDNA) sequences, and (3) have DNA helix-destabilizing ("unwinding") activity. We have characterized both dsDNA binding and helix destabilization. The affinity of RPA for dsDNA was lower than that of ssDNA and precisely correlated with the melting temperature of the DNA fragment. The rates of helix destabilization and dsDNA binding were similar, and both were slow relative to the rate of binding ssDNA. We have previously mapped the regions required for ssDNA binding [Walther et al. (1999) Biochemistry 38, 3963-3973]. Here, we show that both helix-destabilization and dsDNA-binding activities map to the central DNA-binding domain of the 70-kDa subunit and that other domains of RPA are needed for optimal activity. We conclude that all types of RPA binding are manifestations of RPA ssDNA-binding activity and that dsDNA binding occurs when RPA destabilizes a region of dsDNA and binds to the resulting ssDNA. The 70-kDa subunit of all RPA homologues contains a highly conserved putative (C-X2-C-X13-C-X2-C) zinc finger. This motif directly interacts with DNA and contributes to dsDNA-binding/unwinding activity. Evidence is presented that a metal ion is required for the function of the zinc-finger motif.

DNA↗
PubMed1997

Role of protein-protein interactions in the function of replication protein A (RPA): RPA modulates the activity of DNA polymerase alpha by multiple mechanisms.

Replication Protein A (RPA) from human cells is a stable complex of 70-, 32-, and 14-kDa subunits that is required for multiple processes in DNA metabolism. RPA binds with high affinity to single-stranded DNA and interacts with multiple proteins, including proteins required for the initiation of SV40 DNA replication, DNA polymerase alpha and SV40 large T antigen. We have used a series of mutant derivatives of RPA to map the regions of RPA required for specific protein-protein interactions and have examined the roles of these interactions in DNA replication. T antigen, DNA polymerase alpha and the activation domain of VP16 all have overlapping sites of interaction in the N-terminal half (residues 1-327) of the 70-kDa subunit of RPA. In addition, the interaction site for DNA polymerase alpha is composed of two functionally distinct regions, one (residues 1- approximately 170) which stimulates polymerase activity and a second (residues approximately 170-327) which increases polymerase processivity. In the latter, both the direct protein-protein interaction and ssDNA-binding activities of RPA were needed for RPA to modulate polymerase processivity. We also found that SV40 T antigen inhibited the ability of RPA to increase processivity of DNA polymerase alpha, suggesting that this activity of RPA may be important for elongation but not during the initiation of DNA replication. DNA polymerase alpha, but not T antigen also interacted with the 32- and/or 14-kDa subunits of RPA, but these interactions did not seem to effect polymerase activity.

Antigens, Polyomavirus Transforming↗
PubMed1994

Vascular architecture of the human optic chiasma and bitemporal hemianopia.

The blood supply system of the optic chiasma was studied in 85 fresh human specimens using various histological and anatomical methods. Computer image analysis and ultrastructural examinations of the microvessel in 58 fetal specimens were also conducted. The authors found that the medial portion of the chiasma is a weak point in the microcirculation network. This weak point is apt to be disturbed first and become ischemic, causing disorders of the crossing optic nerve fibers and resulting in characteristic bitemporal visual field defects. SEM studies showed no ultrastructural difference between the capillaries at the medial and lateral portions of the chiasma. It was concluded that: 1) No special artery supplies the median chiasma the weak point of microcirculation at the median chiasma is due to its relatively scanty capillary distribution; 2) "lateral chiasma arteries" could provide a better blood supply to the lateral fibers and thus the nasal quadrantic visual field could be preserved in many late stages of visual field defect in sellar region tumors; 3) cases with pituitary microadenoma which is not sufficiently large to press the chiasma but involves bitemporal visual field defect are due to the tumor receiving "shunt-flow" (stealing blood) from the chiasma through the peri-infundibulum plexus.

Capillaries↗