Pur-alpha is a protein that in humans is encoded by the PURAgene[1] located at chromosome 5, band q31.[2][3]
Pur-alpha an ancient, multi-functional DNA- and RNA-binding protein.[1][4]PURA is expressed in every human tissue. Human Pur-alpha is a protein of 322 amino acids. According to convention, PURA, the gene, is written italicized in all upper case letters. Pur-alpha, the protein, is written with the first letter capitalized and can be found listed as Pur-alpha, Pur-α, Pura, Puralpha, Pur alpha and Pur1.
Pur-alpha was the first sequence-specific single-stranded DNA-binding protein to be discovered in higher organisms (GenBank M96684.1; GI:190749).[1] It binds to both single-stranded and double-stranded DNA, making contact with G residues in the purine-rich strand of its binding site. Cumulative data shows that Pur-alpha preferentially binds to the sequence (G2-4N1-3)n, where N is not G. N denotes a nucleotide, and n denotes the number of repeats of this small sequence. N may be repeated up to three times in this sequence.[1][5] Following the identification of a Pur factor, which specifically bound a purine-rich sequence in the control region of the c-MYC gene,[6] the gene, PURA, encoding the protein, Pur-alpha, was cloned and sequenced for both human[1] and mouse (GenBank U02098.1).[4] Pur-alpha belongs to the four-member Pur protein family, which also includes Pur-beta (GenBank AY039216.1; GI:14906267)[1] and two forms of Pur–gamma (Variant A, GenBank AF195513.2; Variant B, GenBank AY077841).[7]
Pur protein sequences from bacteria through humans contain an amino acid segment that is strongly conserved (see NCBI smart00712).[1][8] Human Pur-alpha contains three repeats of this Pur domain and bacterial Pur-alpha contains one.[1][9] This evolutionary conservation means that the specific sequence of this domain is important for the survival of most species throughout the spectrum of living organisms. This essential nature of the Pur domain piques interest because the functions of Pur-alpha in lower organisms and in humans differ greatly. For example, Pur-alpha is essential for brain and blood cell development in mammals,[10] but bacteria have no brain and no blood. In humans Pur-alpha functions to activate transcription in the nucleus, to facilitate RNA transport in the cytoplasm and to regulate DNA replication in the cell cycle.[8] In certain functions Pur-alpha interacts with family member Pur-beta.[11][12] Several cell cycle regulatory functions may be mediated by Pur-alpha binding to Cyclin/Cdk protein kinases, which phosphorylate proteins regulating cell cycle transition points.[13][14] Requirements for Pur-alpha in all organisms are united by Pur-alpha’s ability to bind nucleic acids coupled to its ability to interact with regulatory and transport proteins.
Relevance in human diseases
Genetic perturbation in leukemia and anti-proliferative effect
PURA, located at chromosome 5 band q31, is frequently deleted in myelodysplastic syndrome (MDS),[15] a disorder of white blood cells, that may progress to acute myelogenous leukemia (AML).[2] Loss of one copy of chromosome 7 is also frequent in MDS. PURB, the gene encoding Pur-beta, is located at 7p13. A visual fluorescence analysis of chromosomes from MDS patients shows that deletions of PURA at 5q31 are more strongly linked to progression of MDS to AML when combined with deletions of the PURB gene, including complete loss of chromosome 7.[2] All of the PURA deletions noted, involve only one of the two paired, parentally-derived chromosomes. The implication is that Pur-alpha and -beta are each codominantly expressed, and that haploid levels are insufficient for a protective effect against cancer. All known PURA deletions in people occur in only one of the two copies of chromosome 5.[16]
Inducing increased levels of Pur-alpha in several different cultured cancer cell lines blocks cell proliferation. It also blocks anchorage-independent colony formation, a hallmark of cancer.[13][17] This is true whether Pur-alpha is microinjected or expressed after introducing a cloned PURA cDNA into cells.[18] The Pur-alpha inhibition of cancer cell proliferation occurs at specific points in the cell division cycle, primarily at checkpoints for transition to DNA replication or mitosis.[18] These cell cycle effects are consistent with an interaction between Pur-alpha and CDK, cell cycle-dependent protein kinases.[13] They are also consistent with documented interaction between Pur-alpha and the tumor suppressor protein, Rb.[19]
Role in mammalian brain development and neurological diseases
Studies of genetic inactivation of PURA in the mouse provided evidence leading to that for PURA gene disorders in brain disease. Homozygous PURA knockouts die shortly after birth with severe defects in brain layer development, tissue wasting and movement disorders. Defects in blood cell development are also prominent, and it is not known how these may affect the brain. Heterozygous knockouts do not die early but exhibit seizure-like disorders.[10] In rat hippocampal neurons, Pur-alpha is found in the cytoplasm together with mRNA transcripts, in a complex including non-coding RNAs, Pur-beta, fragile X mental retardation proteins and microtubule-associated proteins. This complex is transported by a kinesin motor[20][21] to sites of translation at junctions of nerve cell dendrites.[22] Recently PURA mutations have been found in multiple patients with brain disorders of a similar phenotype including hypotonia, developmental delay, movement disorders, and seizure or seizure-like movements.[23][24][25] This spectrum of brain disorders is similar to the phenotype of a central nervous system syndrome termed the 5q31.3 microdeletion syndrome,[23] and is the basis for a proposed PURA Syndrome[26] based on PURA mutations rather than just deletions.
Influence on HIV-1 replication
In the brain Pur-alpha plays a role in diseases involving glial cells, cells that support nerve cells, as well as diseases involving nerve cells. These diseases include neuro-AIDS. Pur-alpha binds to a regulatory RNA element, called TAR, in the HIV-1 genome.[27] This activates the expression of Tat, a transcriptional activator of its own gene. Pur-alpha binds TAR, allowing Tat to bind an adjacent TAR site to stimulate transcription. Pur-alpha then binds to the Tat protein itself. Pur-alpha also binds Cyclin T1, a regulatory partner of Cdk9 protein kinase, necessary for Tat activity. Cyclin T1/Cdk9 phosphorylates a region of RNA polymerase II. Such phosphorylation of the polymerase enhances its ability to complete RNA synthesis and stimulates replication of the HIV-1 RNA genome.[28][29]
Cooperative effect with HIV-1 on JC polyomavirus replication and expression
Pur-alpha participates in development of progressive multifocal leukoencephalopathy (PML), a loss of the nerve sheath formed by oligodendroglial cells.[30][31][28] Although HIV-1 is not usually found in these glial cells, HIV-1 proteins can pass through cell membranes to enter them. JCV is considered the causative agent of PML. JCV is activated in the glial cells by certain states of immune system suppression, including HIV-1 infection.[32] There is a documented interaction between Pur-alpha, the HIV-1 protein, Tat, and a Pur-alpha-binding regulatory sequence in JCV DNA.[31] Pur-alpha acts by altering both replication and gene expression of JCV.[30][33][34][31][35]
Role in amyotrophic lateral sclerosis (ALS)
Pur-alpha plays a role in ALS, otherwise known as Lou Gehrig’s disease. ALS is a motor neuron disease involving both the brain and spinal cord, resulting in progressive loss of muscle control. ALS has several contributing causes, but the most common familial form is due to an expanded repeat of the hexanucleotide GGGGCC at the chromosomal locus C9ORF72.[36][37] The C9ORF72 hexanucleotide repeat expansion (HRE) is capable of binding Pur-alpha very tightly. Pur-alpha may act in ALS directly by binding this DNA repeat expansion or its single-stranded RNA transcript.[38][37] One potential consequence of this binding would be to influence an unconventional translation of this transcript repeat that results in long dipeptide repeats. This is termed RAN (Repeat Associated Non-ATG) translation initiation.[39] Aberrant Pur-alpha association with its RNA sequence segment may also be a feature of ALS types that do not involve C9ORF72 expansion.[40] Addition of Pur-alpha suppresses neurodegeneration in mouse neuronal cells and in Drosophila expressing the C9ORF72 HRE.[37] Pur-alpha also reverses neuronal changes caused by defects in the gene, FUS, which can lead to ALS.[40][41] The mechanism of action of Pur-alpha in ALS is not known. There is presently no evidence that the PURA sequence itself is mutated in the C9ORF72 form of ALS. Rather, it is a regulatory nucleic acid sequence to which Pur-alpha binds that is altered.
↑ 2.02.12.2Lezon-Geyda K, Najfeld V, Johnson EM (June 2001). "Deletions of PURA, at 5q31, and PURB, at 7p13, in myelodysplastic syndrome and progression to acute myelogenous leukemia". Leukemia. 15 (6): 954–62. PMID11417483.
↑Ma ZW, Pejovic T, Najfeld V, Ward DC, Johnson EM (1995). "Localization of PURA, the gene encoding the sequence-specific single-stranded-DNA-binding protein Pur alpha, to chromosome band 5q31". Cytogenetics and Cell Genetics. 71 (1): 64–7. PMID7606931.
↑ 4.04.1Ma ZW, Bergemann AD, Johnson EM (November 1994). "Conservation in human and mouse Pur alpha of a motif common to several proteins involved in initiation of DNA replication". Gene. 149 (2): 311–4. PMID7959008.
↑Wortman MJ, Johnson EM, Bergemann AD (March 2005). "Mechanism of DNA binding and localized strand separation by Pur alpha and comparison with Pur family member, Pur beta". Biochimica et Biophysica Acta. 1743 (1–2): 64–78. doi:10.1016/j.bbamcr.2004.08.010. PMID15777841.
↑ 8.08.1Johnson EM (2003). "The Pur protein family: clues to function from recent studies on cancer and AIDS". Anticancer Research. 23 (3A): 2093–100. PMID12894583.
↑Hariharan S, Kelm RJ, Strauch AR (September 2014). "The Purα/Purβ single-strand DNA-binding proteins attenuate smooth-muscle actin gene transactivation in myofibroblasts". Journal of Cellular Physiology. 229 (9): 1256–71. doi:10.1002/jcp.24564. PMID24446247.
↑Kelm RJ, Elder PK, Strauch AR, Getz MJ (October 1997). "Sequence of cDNAs encoding components of vascular actin single-stranded DNA-binding factor 2 establish identity to Puralpha and Purbeta". The Journal of Biological Chemistry. 272 (42): 26727–33. PMID9334258.
↑ 13.013.113.2Barr SM, Johnson EM (2001). "Ras-induced colony formation and anchorage-independent growth inhibited by elevated expression of Puralpha in NIH3T3 cells". Journal of Cellular Biochemistry. 81 (4): 621–38. PMID11329617.
↑Liu H, Barr SM, Chu C, Kohtz DS, Kinoshita Y, Johnson EM (March 2005). "Functional interaction of Puralpha with the Cdk2 moiety of cyclin A/Cdk2". Biochemical and Biophysical Research Communications. 328 (4): 851–7. doi:10.1016/j.bbrc.2005.01.038. PMID15707957.
↑Reference, Genetics Home. "PURA gene". Genetics Home Reference.
↑Hirai, H (April 2003). "Molecular mechanisms of myelodysplastic syndrome". Japanese journal of clinical oncology. 33 (4): 153–60. PMID12810828.
↑Darbinian N, Gallia GL, King J, Del Valle L, Johnson EM, Khalili K (December 2001). "Growth inhibition of glioblastoma cells by human Pur(alpha)". Journal of Cellular Physiology. 189 (3): 334–40. doi:10.1002/jcp.10029. PMID11748591.
↑ 18.018.1Stacey DW, Hitomi M, Kanovsky M, Gan L, Johnson EM (July 1999). "Cell cycle arrest and morphological alterations following microinjection of NIH3T3 cells with Pur alpha". Oncogene. 18 (29): 4254–61. doi:10.1038/sj.onc.1202795. PMID10435638.
↑Johnson EM, Chen PL, Krachmarov CP, Barr SM, Kanovsky M, Ma ZW, Lee WH (October 1995). "Association of human Pur alpha with the retinoblastoma protein, Rb, regulates binding to the single-stranded DNA Pur alpha recognition element". The Journal of Biological Chemistry. 270 (41): 24352–60. PMID7592647.
↑Kanai Y, Dohmae N, Hirokawa N (August 2004). "Kinesin transports RNA: isolation and characterization of an RNA-transporting granule". Neuron. 43 (4): 513–25. doi:10.1016/j.neuron.2004.07.022. PMID15312650.
↑Kobayashi S, Agui K, Kamo S, Li Y, Anzai K (October 2000). "Neural BC1 RNA associates with pur alpha, a single-stranded DNA and RNA binding protein, which is involved in the transcription of the BC1 RNA gene". Biochemical and Biophysical Research Communications. 277 (2): 341–7. doi:10.1006/bbrc.2000.3683. PMID11032728.
↑Johnson EM, Kinoshita Y, Weinreb DB, Wortman MJ, Simon R, Khalili K, Winckler B, Gordon J (May 2006). "Role of Pur alpha in targeting mRNA to sites of translation in hippocampal neuronal dendrites". Journal of Neuroscience Research. 83 (6): 929–43. doi:10.1002/jnr.20806. PMID16511857.
↑Berger, JR; Chauhan, A; Galey, D; Nath, A (August 2001). "Epidemiological evidence and molecular basis of interactions between HIV and JC virus". Journal of NeuroVirology. 7 (4): 329–38. PMID11517412.
↑Daniel DC, Kinoshita Y, Khan MA, Del Valle L, Khalili K, Rappaport J, Johnson EM (December 2004). "Internalization of exogenous human immunodeficiency virus-1 protein, Tat, by KG-1 oligodendroglioma cells followed by stimulation of DNA replication initiated at the JC virus origin". DNA and Cell Biology. 23 (12): 858–67. doi:10.1089/dna.2004.23.858. PMID15684713.
↑Daniel DC, Wortman MJ, Schiller RJ, Liu H, Gan L, Mellen JS, Chang CF, Gallia GL, Rappaport J, Khalili K, Johnson EM (July 2001). "Coordinate effects of human immunodeficiency virus type 1 protein Tat and cellular protein Puralpha on DNA replication initiated at the JC virus origin". The Journal of General Virology. 82 (Pt 7): 1543–53. doi:10.1099/0022-1317-82-7-1543. PMID11413364.
↑Wortman MJ, Krachmarov CP, Kim JH, Gordon RG, Chepenik LG, Brady JN, Gallia GL, Khalili K, Johnson EM (February 2000). "Interaction of HIV-1 Tat with Puralpha in nuclei of human glial cells: characterization of RNA-mediated protein-protein binding". Journal of Cellular Biochemistry. 77 (1): 65–74. PMID10679817.
↑Rossi S, Serrano A, Gerbino V, Giorgi A, Di Francesco L, Nencini M, et al. (May 2015). "Nuclear accumulation of mRNAs underlies G4C2-repeat-induced translational repression in a cellular model of C9orf72 ALS". Journal of Cell Science. 128 (9): 1787–99. doi:10.1242/jcs.165332. PMID25788698.
Barbe MF, Krueger JJ, Loomis R, Otte J, Gordon J (November 2016). "Memory deficits, gait ataxia and neuronal loss in the hippocampus and cerebellum in mice that are heterozygous for Pur-alpha". Neuroscience. 337: 177–190. doi:10.1016/j.neuroscience.2016.09.018. PMID27651147.
Weber J, Bao H, Hartlmuller C, Wang Z, Windhager A, Janowski R, Madl T, Jin P, Niessing D (2016). "Structural basis of nucleic acid recognition and double-strand unwinding by the essential neuronal protein Pur-alpha". Elife. 5. doi:10.7554/eLife.11297. PMID26744780.
Peruzzi F, Gordon J, Darbinian N, Amini S (December 2002). "Tat-induced deregulation of neuronal differentiation and survival by nerve growth factor pathway". Journal of Neurovirology. 8 Suppl 2 (2): 91–6. doi:10.1080/13550280290167885. PMID12491158.
Tretiakova A, Steplewski A, Johnson EM, Khalili K, Amini S (October 1999). "Regulation of myelin basic protein gene transcription by Sp1 and Puralpha: evidence for association of Sp1 and Puralpha in brain". Journal of Cellular Physiology. 181 (1): 160–8. doi:10.1002/(SICI)1097-4652(199910)181:1<160::AID-JCP17>3.0.CO;2-H. PMID10457364.
Darbinian N, Gallia GL, Kundu M, Shcherbik N, Tretiakova A, Giordano A, Khalili K (November 1999). "Association of Pur alpha and E2F-1 suppresses transcriptional activity of E2F-1". Oncogene. 18 (46): 6398–402. doi:10.1038/sj.onc.1203011. PMID10597240.
Itoh H, Wortman MJ, Kanovsky M, Uson RR, Gordon RE, Alfano N, Johnson EM (August 1998). "Alterations in Pur(alpha) levels and intracellular localization in the CV-1 cell cycle". Cell Growth & Differentiation. 9 (8): 651–65. PMID9716182.
Gallia GL, Safak M, Khalili K (December 1998). "Interaction of the single-stranded DNA-binding protein Puralpha with the human polyomavirus JC virus early protein T-antigen". The Journal of Biological Chemistry. 273 (49): 32662–9. doi:10.1074/jbc.273.49.32662. PMID9830007.
Thatikunta P, Sawaya BE, Denisova L, Cole C, Yusibova G, Johnson EM, Khalili K, Amini S (December 1997). "Identification of a cellular protein that binds to Tat-responsive element of TGF beta-1 promoter in glial cells". Journal of Cellular Biochemistry. 67 (4): 466–77. doi:10.1002/(SICI)1097-4644(19971215)67:4<466::AID-JCB5>3.0.CO;2-Q. PMID9383706.
