Manual Emilia Gainey

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As it opens over time the richness becomes more pronounced and the texture Choose Options Quick view. Julius was the son of Septimus and Mary Tyler Huntington. He was also married to Margaret Gainey. I am interested in finding out as much information as possible about Julius and his wife Marg Gainey are buried at Pleasant Ridge Cemetery in I don't know for certain the death location of Roxanna Rehberg presumably that is her name as it appears on her grave stone Simon Brewer, Wilkinson Co.

This Daniel married Martha Gainey and they all lived in the same area near the town of Gordon. Daniel was about the same age a Re: Cloice W. Edward W. Brooks, not C. If anyone is doing research on this particular line, I have information to share. Ewel W. Barker on Jan 11, ; and Jane Dagley m. New Gainey Website. We will shortly be uploading a new website for TheGaineyfamily. Please send the addresses to admin theGaineyfamily. I spoke to my mother who is a Gainey , she did not know your grandfather but had heard of him.

She doesn't know for sure if your grandfather was related, but said it is a good chance. She said he lived in no Any information would be greatly appreciated! Tom, do you know any of the Whittingtons who still live in Marion, SC? I'm trying to locate the family of Eugene "Tootsie" Whittington who was probably born in the early 's. If anyone would like my updated Ratledge file e-mail me at ddeskin Hi Darren, Just came from the message board.

Would it be possible to a copy of the info that Houston Ratledge sent to you? Also could you please scan or make copies will be more then glad to pay an Ratledge who married Ruth. Here, we identify determinants of nuclear cGAS localization and activation. The non-enzymatic N-terminal domain of cGAS determines nucleo-cytoplasmic localization, enrichment on centromeres, and activation of nuclear-localized cGAS. These results reveal a preferential functional association of nuclear cGAS with centromeres. While mitochondrial integrity is linked to cell survival and its disruption leads to apoptotic cell death Tait and Green, , the nuclear envelope NE is a dynamic barrier in both cycling or differentiated non-dividing cells.

Nuclear disassembly leaves the nuclear DNA potentially accessible to cytosolic factors. Moreover, the confinement of interphase cells, such as during migration in tissues, leads to repeated nuclear envelope rupture and repair events. We asked how cGAS recruitment to the nucleus is determined and to what extent cGAS could be activated in the nucleus itself. C Nuclear-cytoplasmic fractionation of post-mitotic human DCs and immunoblots for endogenous cGAS top , tubulin center , and lamin B1 bottom.

We hypothesized that endogenous nuclear cGAS in interphase could result from the interaction with nuclear DNA during nuclear envelope breakdown that occurred in a previous mitosis. At early metaphase, cGAS started to accumulate at the periphery of the nucleus, coinciding with the onset of nuclear envelope breakdown. After nuclear envelope breakdown and through mitosis, cGAS accumulated on distinctive chromosomes. Therefore, cGAS is expressed as a cytosolic protein in interphase, but nuclear envelope breakdown during the cell cycle renders the DNA available for cGAS binding and recruitment in the nucleus, and this generates daughter cells with a pool of nuclear cGAS that persists during the next interphase.

One cell is going through mitosis. Red indicates nuclear mask. Blue corona indicates cytoplasmic mask. Various cells are going through mitosis. Time is hh:mm. We aimed to test whether increased levels of nuclear cGAS could lead to the innate activation of DCs. We measured the induction of the co-stimulatory molecule CD86, a marker of innate immune activation in DCs.


These results indicate that increasing the nuclear cGAS level in DCs results in innate immune activation. We concluded that nuclear-localized cGAS is enzymatically active and limited by at least fold compared to its activity in the response to transfected DNA. Dilutions are 3-fold. One-sample t test. Gray dashed line indicates lower limit of detection; bar shows geometric mean.

Gray dashed line indicates lower limit of detection. Bar shows geometric mean. One representative field. The cells were confined and the movie was started after compression. NE rupture events increase during time. Cells were transfected and the movie was started. To determine the enrichment of cGAS on genomic features, we computed the fraction of peaks falling within a given genomic element compared to the expected fraction based on the genomic coverage. We reasoned that cGAS may be associated with a subset of specific satellite occurrences.

To account for the differences in the number of individual repeat elements between classes, we sorted the elements by enrichment and computed the fraction of occurrences within rank bins within each class. The cGAS track represents the fold change chip over input of selected filtered peaks regions from the 3 donors. The CENP-B box track reports on the x axis the genomic position of the region occurrence of CENP-B box consensus sequence and on the y axis the minimal distance log10 transformed of the region to its two neighboring regions.

G cGAS-specific read enrichment on repeats. Means and SDs are represented.

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We wondered whether the centromeric association of cGAS was determined in the protein sequence. To understand whether the N-terminal domain could be a functional determinant in cGAS, we expressed the truncated domains in non-cycling human DCs. We next examined the intracellular localization of the isolated domain 1— We conclude that cGAS expressed in interphase is actively retained in the cytosol by domain 1—, which counteracts two nuclear-localizing activities in domains 1— and — GFP channel is shown in black on white.

Bars represent geometric means. We hypothesized that domain 1— may determine the association of nuclear cGAS with centromeres. These results show that once cGAS is in the nucleus, the N-terminal domain 1— is required for association with centromeric DNA and activation of the sensor. Finally, we sought to determine whether centromere association also applied to endogenous cGAS.

First, we stained endogenous cGAS on metaphase spread chromosomes in a cycling human cell line. In contrast to human centromeres, mouse centromeres are telocentric and poorly mapped in the reference genome. In particular, minor satellites, which constitute mouse centromeres, are not annotated on mm A Representative immunofluorescence images of a metaphase spread of cycling U2OS cells showing endogenous cGAS enrichment at the inner centromere.

CENP-A marks the centromere position. Yellow arrows point to cGAS localization at an inner centromere. Error bars represent the SEMs of 36 centromeres in 1 cell representative of 2 independent experiments.

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Bars represent means and error bars are SEMs. J Working model of cGAS expression in the cytoplasm in interphase, followed by localization to the nucleus as a result of mitosis.

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  • We provided four distinct pieces of evidence that support this finding. Third, endogenous cGAS was directly observed on the centromeres of metaphase chromosomes. We also show that the N-terminal domain of cGAS, which is dispensable for the catalytic activity of the recombinant protein and the response to transfected DNA, demonstrates distinct activities according to cGAS localization. When cGAS is cytosolic in interphase, N-terminal domain 1— encodes a dominant cytosolic retention activity.

    When this domain is disrupted in the isolated cGAS fragments 1— and —, the presence of nuclear-localization signals is revealed in both fragments. When cGAS is nuclear, domain 1— is required for association with centromeric DNA and for innate immune activation by the nuclear-localized sensor.

    Whether this enhancement is functionally linked to subcellular localization remains to be determined. In accordance with this hypothesis, we find that transfected four-repeat satellite DNA fragments induce a stronger cellular innate immune activation compared to shuffled sequence, and this difference is lost with increasing numbers of repeats. These findings require further study, and we speculate that additional types of chromosomal DNA contribute to the regulation of nuclear cGAS activity.

    It will be important to develop assays to determine the contribution of DNA sequences and chromatin proteins in regulating cGAS enzymatic activity for centromeric and non-centromeric DNA. Since monocyte-derived DCs do not divide, our results indicate that endogenous cGAS is maintained in the nucleus for several days. This could result from a sustained retention of endogenous cGAS in the nucleus, or alternatively from the replenishment of nuclear cGAS during interphase.

    We find that nuclear-localized cGAS functionally upregulates cellular innate immune responses. Alternatively, we do not exclude that a fraction of cytosolic cGAMP detected in the experiment originated from cGAMP contained in the lentiviral particles that were used for DC transduction.

    Our data show that nuclear cGAS activity is restrained by at least four mechanisms. First, overexpressing nuclear-localized cGAS activates innate immunity in DCs, suggesting that the endogenous level of expression of the sensor in DCs is tuned to avoid spontaneous activation. This suggests that enzymatic activation in the nucleus is limited by a yet-to-be-elucidated mechanism. Third, the N-terminal domain of cGAS is crucial to retain the sensor in the cytosol until a nuclear envelope rupture or nuclear envelope breakdown occurs.

    Fourth, where cGAS interacts with nuclear DNA appears to determine cGAS activation: while cGAS is activated after nuclear entry resulting from mitosis or association with a nuclear-localization signal when the sensor is overexpressed, we could not detect endogenous cGAS activation after entry through interphasic nuclear envelope rupture events. In addition, nuclear cGAS may be endowed with nuclear-specific functions that future work may unveil. Further information and requests for reagents may be directed to and will be fulfilled by the Lead Contact, Nicolas Manel nicolas.

    Healthy individuals from Paris area donate venous blood to be used for research. Gender identity and age from anonymous healthy donors was not available. EFS provides informed consent to blood donors.

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    Cell lines are described in the Key Resources Table. Number of experimental replicates are indicated in the respective figure legends. Number of donors and experimental replicates are indicated in the respective figure legends. All animal procedures were in accordance with the guidelines and regulations of the French Veterinary Department in an accredited animal facility.

    Mice used in experiments were females. All mice in each experiment were littermates. Mouse bone marrow derived DCs were differentiated from bone marrow isolated from mouse tibiae. Cells were split at day 4 and day 7 and harvested at day Cells were counted and reseeded in BMDCs medium at a concentration of 0.