KLRG1 Overexpression Lysate

Western Blot: KLRG1 Overexpression Lysate (Adult Normal) [NBP2-07698] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for KLRG1.

• Reactivity: Hu
• Applications: WB

KLRG1 Overexpression Lysate Summary

• Description:

  KLRG1 Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC219705
  
  Accession#: NM_005810
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: KLRG1

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
• Theoretical MW: 21 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for KLRG1 Overexpression Lysate

• 2F1
• CLEC15A
• CLEC15AMAFA-LIKE
• C-type lectin domain family 15 member A
• C-type lectin domain family 15, member A
• ITIM-containing receptor MAFA-L
• killer cell lectin-like receptor subfamily G member 1
• killer cell lectin-like receptor subfamily G, member 1
• KLRG1
• MAFA
• MAFAL
• MAFA-L
• MAFA-like receptor
• MAFAMAFA-2F1
• mast cell function-associated antigen (ITIM-containing)
• Mast cell function-associated antigen
• MGC13600

Background

KLRG1 (killer cell lectin-like receptor G1), also known as CLEC15A and MAFA, belongs to the KLR family. KLRG1 is a lectin-like inhibitory receptor that is a single pass type II transmembrane protein (1). KLRG1 functions in innate immunity and has an inhibitory role on the function of natural killer (NK) cells and T cells that is induced by binding to their non-MHC ligands (1,2). The KLRG1 protein exists as both a monomer and homodimer and is expressed as an inhibitory receptor on NK cells as well as subsets of CD4+ and CD8+ T cells, gammadelta T cells, and alphabeta T cells (1,3,4). The human KLRG1 protein is 195 amino acids (aa) in length with a theoretical molecular weight (MW) of 21.8 kDa (2). Given KLRG1 protein glycosylation and disulfide bonds, the observed molecular weight often ranges from 30-38 kDa. The human KLRG1 protein consists of a cytoplasmic domain at 1-38 aa with one immunoreceptor tyrosine-based inhibitory motif (ITIM) at 5-10 aa, a transmembrane segment at 39-59 aa, and an extracellular domain (ECD) at 60-195 aa with one C-type lectin domain (CTLD) at 82-185 aa (2).

KLRG1's primary ligand is E-cadherin, which is expressed on non-neural epithelial tissues, but the KLRG1 receptor also recognizes both N- and R-cadherin expressed in nervous system tissues (1,3,5,6). E-cadherin binding of KLRG1 leads to ITIM triggering and, ultimately, either reduced T cell proliferation or decreased NK cell cytotoxicity (3,5,6). More precisely, upon differentiation of CD8+ T cells, cells switch from signaling through CD28 and instead signal through the inhibitory KLRG1 receptor molecule (3,4). Phosphorylation of KLRG1's ITIM causes recruitment of two phosphatases, SH2-containing inositol polyphosphate 5-phoshate (SHIP-1) and SH-2 containing protein-tyrosine phosphatase 2 (SHP-2) (3,5). The effectors SHIP-1 and SHP-2 degrade PIP3 to PIP2, preventing Akt phosphorylation and inhibiting proliferation (3,5,6). Conversely, when KLRG1 signaling is blocked in highly differentiated T cells, Akt signaling is restored and cell proliferation resumes (3). An increase in highly differentiated T cells is observed in many age-related infections such as meningitis, pneumonia, and influenza, which also correlates with elevated KLRG1 levels (3). This observation suggests that KLRG1 may be a potential immunotherapeutic target, especially for vaccinations which typically have decreased response in aged patients (3).

References

1. Li Y, Hofmann M, Wang Q, Teng L, Chlewicki LK, Pircher H, Mariuzza RA. Structure of natural killer cell receptor KLRG1 bound to E-cadherin reveals basis for MHC-independent missing self recognition. Immunity. 2009 Jul 17;31(1):35-46. http://doi.org/10.1016/j.immuni.2009.04.019

2. Uniprot(Q96E93)

3. Henson SM, Akbar AN. KLRG1--more than a marker for T cell senescence. Age (Dordr). 2009 Dec;31(4):285-91. http://doi.org/10.1007/s11357-009-9100-9.

4. Thimme R, Appay V, Koschella M, Panther E, Roth E, Hislop AD, Rickinson AB, Rowland-Jones SL, Blum HE, Pircher H. Increased expression of the NK cell receptor KLRG1 by virus-specific CD8 T cells during persistent antigen stimulation. J Virol. 2005 Sep;79(18):12112-6.http://doi.org/10.1128/JVI.79.18.12112-12116.2005.

5. Borys SM, Bag AK, Brossay L, Adeegbe DO. The Yin and Yang of Targeting KLRG1+ Tregs and Effector Cells. Front Immunol. 2022 Apr 29;13:894508. http://doi.org10.3389/fimmu.2022.894508.

6. Van den Bossche J, Malissen B, Mantovani A, De Baetselier P, Van Ginderachter JA. Regulation and function of the E-cadherin/catenin complex in cells of the monocyte-macrophage lineage and DCs. Blood. 2012 Feb 16;119(7):1623-33. http://doi.org/10.1182/blood-2011-10-384289.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: KLRG1