Osmium tetroxide footprinting of a scaffold attachment region in the maizeAdh1 promoter

1993 ◽  
Vol 22 (6) ◽  
pp. 1145-1151 ◽  
Author(s):  
Anna-Lisa Paul ◽  
Robert J. Ferl
Keyword(s):  
Osmium Tetroxide ◽  
Attachment Region ◽  
Scaffold Attachment Region

A matrix/scaffold attachment region binding protein: Identification, purification, and mode of binding

Cell ◽  
1991 ◽  
Vol 64 (1) ◽  
pp. 123-135 ◽  
Author(s):  
Jens P. von Kries ◽  
Hartmut Buhrmester ◽  
Wolf H. Strätling
Keyword(s):  
Protein Identification ◽  
Binding Protein ◽  
Attachment Region ◽  
Scaffold Attachment Region

scholarly journals SAF-Box, a Conserved Protein Domain That Specifically Recognizes Scaffold Attachment Region DNA

2000 ◽  
Vol 20 (20) ◽  
pp. 7480-7489 ◽  
Author(s):  
Michael Kipp ◽  
Frank Göhring ◽  
Thorsten Ostendorp ◽  
Cornelis M. van Drunen ◽  
Roel van Driel ◽  
...  
Keyword(s):  
Dna Binding ◽  
High Specificity ◽  
Binding Mode ◽  
Protein Domain ◽  
Nuclear Scaffold ◽  
Evolutionarily Conserved ◽  
Attachment Region ◽  
Dna Elements ◽  
Scaffold Attachment Region ◽  
Different Origins

ABSTRACT SARs (scaffold attachment regions) are candidate DNA elements for partitioning eukaryotic genomes into independent chromatin loops by attaching DNA to proteins of a nuclear scaffold or matrix. The interaction of SARs with the nuclear scaffold is evolutionarily conserved and appears to be due to specific DNA binding proteins that recognize SARs by a mechanism not yet understood. We describe a novel, evolutionarily conserved protein domain that specifically binds to SARs but is not related to SAR binding motifs of other proteins. This domain was first identified in human scaffold attachment factor A (SAF-A) and was thus designated SAF-Box. The SAF-Box is present in many different proteins ranging from yeast to human in origin and appears to be structurally related to a homeodomain. We show here that SAF-Boxes from four different origins, as well as a synthetic SAF-Box peptide, bind to natural and artificial SARs with high specificity. Specific SAR binding of the novel domain is achieved by an unusual mass binding mode, is sensitive to distamycin but not to chromomycin, and displays a clear preference for long DNA fragments. This is the first characterization of a specific SAR binding domain that is conserved throughout evolution and has DNA binding properties that closely resemble that of the unfractionated nuclear scaffold.

Characterization of a Plant Scaffold Attachment Region in a DNA Fragment That Normalizes Transgene Expression in Tobacco

1992 ◽  
Vol 4 (4) ◽  
pp. 463 ◽  
Author(s):  
Peter Breyne ◽  
Marc Van Montagu ◽  
Ann Depicker ◽  
Godelieve Gheysen
Keyword(s):  
Transgene Expression ◽  
Attachment Region ◽  
Dna Fragment ◽  
Scaffold Attachment Region

Performance- and safety-enhanced lentiviral vectors containing the human interferon-β scaffold attachment region and the chicken β-globin insulator

Blood ◽  
2003 ◽  
Vol 101 (12) ◽  
pp. 4717-4724 ◽  
Author(s):  
Ali Ramezani ◽  
Teresa S. Hawley ◽  
Robert G. Hawley
Keyword(s):  
Transgene Expression ◽  
Retroviral Vectors ◽  
Hematopoietic Progenitor Cell ◽  
Human Interferon ◽  
Interferon Β ◽  
Reporter Expression ◽  
Hematopoietic Progenitor ◽  
Human Cd34 ◽  
Attachment Region ◽  
Scaffold Attachment Region

AbstractRetroviral vectors are the most efficient means of stable gene delivery to hematopoietic stem cells (HSCs). However, transgene expression from retroviral vectors is frequently subject to the negative influence of chromosomal sequences flanking the site of integration. Toward the development of autonomous transgene expression cassettes, we inserted the human interferon-β scaffold attachment region (IFN-SAR) and the chicken β-globin 5′ DNase I hypersensitive site 4 (5′HS4) insulator both separately and together into a series of self-inactivating (SIN) lentiviral vector backbones. Transduced cells of the human CD34+ hematopoietic progenitor line KG1a—pooled populations as well as individual clones harboring single integrants—were analyzed for reporter expression during culture periods of up to 4 months. Vectors carrying both the 5′HS4 insulator and the IFN-SAR consistently outperformed control vectors without inserts as well as vectors carrying either element alone. The performance of a set of vectors containing the murine stem cell virus long terminal repeat as an internal promoter was subsequently assessed during in vitro monocytic differentiation of transduced primary human CD34+ cord blood cells. Similar to what was observed in the KG1a hematopoietic progenitor cell model, optimal reporter expression in primary monocytes was obtained with the vector bearing both regulatory elements. These findings indicate that the 5′HS4/IFN-SAR combination is particularly effective at maintaining open chromatin domains permissive for high-level transgene expression at early and late stages of hematopoietic development, and thus could be of utility in HSC-directed retroviral vector–mediated gene transfer applications.

High-Level Transgene Expression in Plant Cells: Effects of a Strong Scaffold Attachment Region from Tobacco

1996 ◽  
Vol 8 (5) ◽  
pp. 899 ◽  
Author(s):  
George C. Allen ◽  
Gerald Hall ◽  
Susan Michalowski ◽  
Winnell Newman ◽  
Steven Spiker ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Plant Cells ◽  
Attachment Region ◽  
Scaffold Attachment Region ◽  
High Level

scholarly journals Human Beta Interferon Scaffold Attachment Region Inhibits De Novo Methylation and Confers Long-Term, Copy Number-Dependent Expression to a Retroviral Vector

2000 ◽  
Vol 74 (6) ◽  
pp. 2671-2678 ◽  
Author(s):  
Qi Dang ◽  
Jennifer Auten ◽  
Ivan Plavec
Keyword(s):  
De Novo ◽  
Retroviral Vector ◽  
Control Vector ◽  
Transfected Cells ◽  
Expression Levels ◽  
Beta Interferon ◽  
Copy Numbers ◽  
Attachment Region ◽  
Scaffold Attachment Region

ABSTRACT Moloney murine leukemia virus-based retroviral vector expression is gradually lost during prolonged in vitro culture of CEMSS T cells. However, when the human beta interferon scaffold attachment region (IFN-SAR) was inserted into the vector immediately upstream of the 3′ long terminal repeat (LTR), expression was maintained for the length of the study (4 months). Clonal analysis of the retrovirus vector-infected CEMSS cells showed that SAR-containing retroviral vector expression levels were positively correlated with the proviral copy numbers (P < 0.0001), while there was no correlation between the proviral copy numbers and expression levels in control vector-infected clones. Thirty-three percent of the CEMSS cell clones infected with the control vector showed evidence of partial or complete methylation in the 5′ LTR region. In sharp contrast, we detected no methylation in the clones infected with the SAR-containing vector. To demonstrate a direct inhibitory effect of methylation on retroviral vector expression, we have transfected 293 cells with in vitro-methylated proviral DNA. In transiently transfected cells, expression of methylated LTR was reduced but not completely inhibited, irrespective of the presence of the IFN-SAR sequence. In stably transfected cells, however, methylation completely abolished expression of the control vector but not of the SAR-containing vector. Furthermore, the expression of the SAR-containing vector was stable over time, indicating the ability of the SAR sequence to alleviate methylation-mediated transcriptional repression of a vector. This study extends our understanding of the mechanisms of retroviral vector inactivation by methylation and provides insight into a functional role for the SAR elements.

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