Application Highlight: Recent uses of TERF2 in immunofluorescence (IF)

Mon, 07/25/2016 - 14:49

Telomeres are a region of repeat nucleotide sequences located at the end of chromosomes to protect our DNA from becoming damaged via end-to-end fusion.  TERF2, or telomeric-repeat binding factor 2, is important for telomere integrity and aids in the formation of the telosome, the telomeric loop, and control of the amount of DNA needed for telomere replication.  When TERF2 is down regulated, an induction of apoptosis and senescence is also witnessed.  TERF2 also interacts with an abundance of proteins in all of the above pathways, including RAP1, Apollo, TOP1, TOP2A and TOP2B.  The visualization of telomeres on chromosomal structures through immunofluorescence (IF) is an important diagnostic tool and is examined more closely in the following research projects using a Novus Biologicals TRF-2 Antibody (NB100-56506).

TRF-2 antibody

TRF-2 Antibody (4A794.15) [NB100-56506] - Staining of TRF2-bound telomeres in human HeLa cells (Courtesy of Fotiadou, et al, 2004).

The first experiment using a TRF-2 Antibody in IF was performed by Xi et al who aimed to further understand the relationship of TERF2 and TERT in cellular proliferation in stem cells and cancer cells alike.  Using a combined approach to genome editing, which includes CRISPR-Cas9 and also an N-Terminal flag tag to TERT, this group established a functional way to visualize TERT in various experimental methods.  Once this procedure was proven, TERT and TERF2 were examined for co-localization across different stages of cell growth in HeLa cells using a TRF-2 Antibody.  Their results showed a small (7%) amount of co-localization between TERT and TERF2 in only the S-phase of cellular growth.  This experiment was the first step in developing a more efficient way to use antibodies and tag sequences side-by-side to examine protein dynamics. 

Next, Yun-Hee Kim et al used a TRF-2 Antibody in their efforts to create a new and advanced method of HCC anticancer treatment achieved by depleting hTERT transcripts with trans-splicing ribozyme.  Specifically, a TRF-2 Antibody was used to show whether contribution by Rz-mediated hTERT depletion after Ad-CRT delivery yielded any significant changes.  After confirming the decrease of hTERT in Ad-CRT treated cells, TERF2 double staining remained normal in Ad-CT infection.  However, after infection with Ad-CRT, telomere staining decreased and TERF2’s overlap with TERF2 was not strong.  To confirm telomere down regulation, they used an antibody for the DNA damage signaling protein 53BP1 and found that this was in fact present. 

Furthermore, Hasegawa et al used a TRF-2 Antibody in IF to investigate a small subpopulation of self-renewable glioma stem cells (GSCs) in glioblastomas that has been implicated in tumor promotion.  While it is known that telomestatin impairs GSC growth, it is unclear exactly how this response correlates with DNA damage.  First, they used a TRF-2 Antibody in IF in GBM146 cells to show that treatment with telomestatin reduced the amount of TERF2 foci, alluding to DNA damage susceptibility.  Interestingly, they found that this TERF2 reduction via telomestatin introduction was faster than the same effect of TERF2 reduction in the presence of the drug Temozolomide, suggesting that there is an affinity from telomestatin for the G4 population of GSC cells.  Overall, these experiments shed light interesting developments regarding the role of TERF2 in this subpopulation of glioma related cells.

Novus Biologicals offers TRF-2 reagents for your research needs including:

  1. PMID: 26553065
  2. PMID: 26909111
  3. PMID: 26845351

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