Tat-Beclin 1: The pioneering of an autophagy-inducing peptide

Wed, 07/20/2016 - 11:39

Autophagy is an essential process that maintains cellular homeostasis and carries out lysosome-mediated degradation of unwanted proteins in the cytoplasm.  Because of this regulatory function, autophagy is often examined when looking at disease pathways.  While our immune system initiates the removal of viruses and pathogens through the autophagic pathway, viruses, such as HIV, have developed a way to evade this process through inhibition.  Therefore, developing a reliable way to examine the molecular process of this inhibition and interaction is very desired.  The central autophagy protein, Beclin 1, became a quick target to create an autophagy manipulation factor and ultimately lead to the formation of an autophagy inducing peptide. Novus Biologicals carries three new isoforms of the Tat-beclin-1 peptide in the form of Tat-Beclin 1 D11 (NBP2-49888), Tat-Beclin 1 L11S (NBP2-49887) and Tat-Beclin L11 (NBP2-49886).  The following articles explain the development and application of these new exciting products. 

tat-beclin 1 peptide

Immunocytochemistry/Immunofluorescence: Tat-Beclin 1 L11 Autophagy Inducing Peptide [NBP2-49886] - HeLa GFP-LC3B cells were treated with Tat-D11, Tat-L11, Tat-Beclin 1 or Tat-L11S for 1.5 hours. Thereafter, the cells were stained using NeuroTrace Red or DAPI and analyzed employing fluorescent microscopy. Note the higher number of autophagosomes/GFP-LC3B+ puncta in the images of Tat-D11 and Tat-L11 treated cells when compared to Tat-Beclin 1 and Tat-L11S treated cells.

Sanae Shoji-Kawata et al are the first to pioneer a candidate therapeutic autophagy-inducing peptide, named "Tat-beclin-1". This peptide can be used alongside primary antibodies, specifically the LC3 antibody, to take a closer look at autophagic dynamics in the cell. To begin, this group started their development knowing that a human immunodeficiency virus (HIV) Nef1 complex exists that interacts with a sequence of the Beclin 1 gene to prevent autophagy.  In order to determine exactly where on the Beclin 1 sequence that Nef1 binds, they performed co-immunoprecipitation assays using FLAG/HA tags on both Beclin1 and Nef and FLAG/HA primary antibodies. Once the amino acid location with the strongest affinity was determined, a cell-permeable peptide coined "Tat-beclin-1" (composed of the HIV-1 Tat protein transduction domain) was developed.  A control peptide, Tat-scrambled, was created simply scrambling the sequence of the Tat-beclin-1 peptide.  Using an LC3 antibody in both western blot and immunofluorescent imaging, it was clear that treatment with the Tat-beclin-1 peptide successfully induced autophagy. 

Next, Sanae Shoji-Kawata et al were interested in which proteins may interact with a biotin conjugated Tat-beclin-1. Through liquid chromatography-tandem mass spectrometry, the golgi-associated plant pathogenesis-related protein 1 (GAPR-1) was identified.  This finding was quite interesting, given that GAPR-1 associated with lipid rafts in the golgi apparatus membrane.  Naturally, once the Tat-beclin-1 peptide had been proven to induce autophagy, the group sought out to examine the potential therapeutic effects of introducing this peptide into a disease model with a need of an autophagic response.  While Tat-beclin-1 did not exhibit a notable effect in vitro in a Huntington’s disease model, it did lessen the clinical effects of neonatal mice infected with the West Nile Virus (WNV) in vivo.  When compared to controls, Tat-beclin-1 peptide treated mice had reduced viral loads in the brain, and subsequently reduced mortality rates.  Overall, it is clear that this peptide has a strong outlook for the field of clinical therapy and research.

Recently, Shirakabe et al used the Tat-beclin-1 peptide along with primary antibodies in their study of Drp1-dependent mitochondrial autophagy in heart failure.  Their findings were similar to the rescue effects of Sanae Shoji-Kawata et al, where they witnessed partial rescue of mitochondrial autophagy and less mitochondrial oxidative stress and heart failure in response to cardiac hypertrophy.  

  1. PMID: 23364696
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  3. PMID: 22257881

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