## Dissertations from ProQuest

#### Title

Transcriptional regulation of the human lambda light chain immunoglobulin locus: Characterization of the 3' enhancer

1994

Article

#### Degree Name

Doctor of Philosophy (Ph.D.)

#### First Committee Member

Bonnie B. Blomberg, Committee Chair

#### Abstract

The first transcriptional enhancer for the human $\lambda$ light chain immunoglobulin locus was recently identified 11.7 kb downstream of C$\lambda$7, the most 3$\sp\prime$ constant region gene. In this study, the cis-acting elements which comprise this enhancer as well as the trans-acting factors which bind to it were examined. By deletion mapping and CAT assays, the complete human $\lambda$ enhancer was determined to reside on a 311 bp PstI-SstI fragment with partial activity retained on a 124 bp PstI-SmaI core element. The complete human $\lambda$ enhancer shares 65% identity with the murine $\lambda$ enhancers, while the core fragment is 70-73% identical. By methylation interference analysis, protein interaction at the well conserved $\lambda{\rm A}$ and $\lambda{\rm B}$ motifs was identified. In addition, protein interaction was observed at a site which is unique to the human $\lambda$ enhancer. This site has been named the Human Enhancer Lambda Protein (HELP) site. From electrophoretic mobility shift assays, a protein factor(s), HELP, has been shown to specifically bind to this site. This motif is located immediately 3$\sp\prime$ of the core enhancer and augments the activity of the core enhancer element. By EMSA analysis, this protein(s) is present primarily in hematopoietic cells but not restricted to B cells. Thus it most likely interacts with another, B cell-restricted protein(s) to play a role in the tissue specific activation of the human $\lambda$ enhancer.The 311 bp human $\lambda$ enhancer is active at the pre-B cell stage, but a larger fragment containing the flanking regions is inactive in pre-B cells whereas it is active in B cells. This suggests the presence of negative elements both upstream and downstream of the complete human $\lambda$ enhancer which are developmentally controlled. By methylation interference analysis, protein interaction at sites in regions 5$\sp\prime$ and 3$\sp\prime$ of the complete human $\lambda$ enhancer were identified which may be responsible for the developmental stage specific activity of this enhancer. The binding site sequence is conserved in both the 5$\sp\prime$ and 3$\sp\prime$ elements, and by cross-competition EMSA the same protein(s) binds to both elements.

#### Keywords

Biology, Molecular; Health Sciences, Immunology