File:Oligomer.mol

2012-01-12T04:54:24Z

Brittany deRonde:

File:Oligomer.pdb

2012-01-12T04:46:05Z

Brittany deRonde:

File:XXX1.mol

2012-01-12T03:57:36Z

Brittany deRonde:

Brittany deRonde: /* Transduction */

One of the [[CBI Molecules]] being studied in the [http://www.umass.edu/cbi/ University of Massachusetts Amherst Chemistry-Biology Interface Program] at UMass Amherst and on display at the [http://www.molecularplayground.org/ Molecular Playground].

== '''Introduction''' ==

'''HIV Tat''', or simply Tat, is a human immunodeficiency virus (HIV) gene that regulates transcription of HIV dsRNA.<ref>Sodroski ''et al. Science.'' '''1985'''. 227, 171-173. [http://www.jstor.org/stable/1695050] http://www.jstor.org/stable/1695050</ref> Tat, which stands for trans-activator of transcription, contains 86 amino acid residues in its sequence.<ref>Arya ''et al. Science'' '''1985'''. 229, 69-73.</ref><ref>Sodroski ''et al. Science.'' '''1985'''. 229, 74-77.</ref> <Structure load='1TIV' size='500' frame='true' align='right' caption='Insert caption here' scene='Brittany_deRonde/Sandbox_1/Hiv_tat/2' /> It is released by HIV infected cells in order to enhance replication of the virus.<ref>Dayton ''et al. Cell.'' '''1986'''. 44, 941-947.</ref><ref>Fisher ''et al. Nature.'' '''1986'''. 320, 367-371.</ref> Nanomolar concentrations of Tat have been reported in the blood of HIV-1 infected people.<ref>Xiao ''et al. Proc. Natl. Acad. Sci. USA.'' '''2000'''. 97, 11466-11471.</ref> When the protein enters non-infected cells, the transcription efficiency increases. It is estimated that transcription levels are 10 to 100 fold higher during the transcription elongation stage when compared to normal basal transcription.<ref>Ensoli ''et al. Nature.'' '''1990'''. 345, 84-86.</ref> Green and Lowenstein, and Frankel and Pabo independently published studies in 1988 that demonstrated that Tat had the ability to cross cellular membranes and initiate transcription of HIV dsRNA.<ref>Green ''et al. Cell.'' '''1988'''. 55, 1179-1188.</ref><ref>Frankel ''et al. Cell.'' '''1988'''. 55, 1189-1193.</ref> These were the first known reports of a '''cell penetrating peptide (CPP)'''.

== '''Cell Penetrating Peptides''' ==

'''Cell penetrating peptides (CPPs)''' are proteins with the ability to cross cellular membranes and facilitate the uptake of various cargo, such as small molecules, siRNA, and small DNA fragments.<ref>Futaki ''et al. J. Biol. Chem.'' '''2001'''. 276, 5836-5840.</ref> Such cargo can be associated via covalent or non-covalent interactions. Tat is considered a CPP because it contains a protein transduction domain (PTD). PTDs are cation-rich sequences of 10-30 residues, usually containing several Lysine and/or Arginine residues.<ref>Wender ''et al. Adv. Drug Deliv. Rev.'' '''2008'''. 60, 452-472.</ref>

[[Image:Arginine1.jpg]] [[Image:Lysine1.jpg]]

These sequences improve interactions between CPPs and cellular membranes and can help them enter cells. The PTD sequence in Tat is YGRKKRRQRRR (amino acid <scene name='Brittany_deRonde/Sandbox_1/Hiv_tat/3'>residues 47-57</scene>), which is Arginine-rich. This 11 amino-acid sequence is now referred to as the TAT peptide, and has be shown to have improved cellular uptake compared to Tat.

Since the discovery of HIV Tat, other Arginine-rich, natural CPPs have been discovered including, penetratin from Drosophila antennapedia protein, sweet arrow peptide (SAP), HIV-1 Rev, flock house virus (FHV) coat, brome mosaic virus (BMV) Gag, human T-cell lymphotrophic virus (HTLV)-II Rex, and the nuclear localization signal (NLS) from nucleoplasmin. <ref>Futaki ''et al. J. Biol. Chem.'' '''2001'''. 276, 5836-5840.</ref><ref>Prochiantz ''et al. Adv. Drug Deliv. Rev.'' '''2008'''. 60, 448-451.</ref> The PTD sequences for these proteins can be found in the table below.

[[Image:Arginine_Rich_Peptides.png]]

== '''Tat-mediated Transduction''' ==

The PTD in HIV Tat is highly cationic. When charged residues (Arginine or Lysine) are replaced with neutral residues (Alanine), cellular uptake decreases. It is likely that the cationic charge is necessary for electrostatic interactions with components of the cellular membrane, such as lipid head groups and proteoglycans.<ref>Ziegler ''et al. Biochemistry'' '''2003'''. 42, 9185-9194.</ref><ref>Goncalves ''et al. Biochemistry.'' '''2005'''. 44, 2692-2702.</ref> Other interactions may also play a role in interactions with the cellular membrane, such as hydrophobic and non-covalent interactions.<ref>Ziegler ''et al. Adv. Drug Deliv. Rev.'' '''2008'''. 60, 580-597.</ref> Although cellular uptake is not directly dependent upon architecture (branched vs. linear), it is dependent upon the number of Arginine residues. Polymers with less than five arginine residues are unable to enter cells.<ref>Wender ''et al. Proc. Natl. Acad. Sci.'' '''2000'''. 97, 13003-13008.</ref> As the number of residues increases, cellular uptake improves up until 15 residues where polymers can enter cells but with hindered efficiencies.<ref>Mitchell ''et al. J. Pept. Res.'' '''2000'''. 56, 318-325.</ref><ref>Futaki ''et al. J. Biol. Chem.'' '''2001'''. 276, 5836-5840.</ref>

==Transduction==
Early studies indicated that HIV Tat primarily entered cells through transduction, meaning that the protein directly crossed the cell membrane. However, for most of these studies, cells were fixed with formaldehyde prior to sample analysis. In 2003, it was discovered that by fixing cells prior to analysis, it permeablized the cell membranes and resulted in artificially high cellular uptake values.

==Endocytosis==

== '''Drug Delivery Applications''' ==

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== '''My Research Interest''' ==

As part of the [http://www.pse.umass.edu/gtew/index.html Tew Research Group] in the [http://www.pse.umass.edu Department of Polymer Science and Engineering] at the University of Massachusetts, Amherst, I am interested in the design of Tat-inspired CPPs for drug and gene delivery. Using functionalized oxanorbornene derivatives as monomers, ring opening metathesis polymerization (ROMP) is performed using Grubbs' 3rd generation catalyst to synthesize cationic CPP mimics. By tuning the functionalization of the monomer structures and the resulting CPP molecular weights, cellular uptake and cell viability can be optimized.

== '''References''' ==
{{Reflist}}