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ВзаимодСйствиС ядСрного Π°ΠΊΡ‚ΠΈΠ½Π° ΠΈ NAP57/дискСрина с Π±Π΅Π»ΠΊΠ°ΠΌΠΈ, спСцифичСски ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΠΌΠΈ ΡΠ°Ρ‚Π΅Π»Π»ΠΈΡ‚Π½ΡƒΡŽ Π”ΠΠš

Π”ΠΈΡΡΠ΅Ρ€Ρ‚Π°Ρ†ΠΈΡΠŸΠΎΠΌΠΎΡ‰ΡŒ Π² Π½Π°ΠΏΠΈΡΠ°Π½ΠΈΠΈΠ£Π·Π½Π°Ρ‚ΡŒ ΡΡ‚ΠΎΠΈΠΌΠΎΡΡ‚ΡŒΠΌΠΎΠ΅ΠΉ Ρ€Π°Π±ΠΎΡ‚Ρ‹

ДостовСрно извСстно, Ρ‡Ρ‚ΠΎ ΡΠ°Ρ‚Π”ΠΠš, ΠΎΠ±Ρ€Π°Π·ΡƒΡŽΡ‰ΠΈΠ΅ Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Ρƒ, ΠΎΠ±Π΅ΡΠΏΠ΅Ρ‡ΠΈΠ²Π°ΡŽΡ‚ Ρ„ΠΎΡ€ΠΌΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΈΠ½Π΅Ρ‚ΠΎΡ…ΠΎΡ€Π°, ΠΊΠΎΡ€Ρ€Π΅ΠΊΡ‚Π½ΠΎΠ΅ спариваниС ΠΈ Ρ€Π°Π·Π΄Π΅Π»Π΅Π½ΠΈΠ΅ хромосом Π² Ρ…ΠΎΠ΄Π΅ митотичСского ΠΈ ΠΌΠ΅ΠΉΠΎΡ‚ичСского Π΄Π΅Π»Π΅Π½ΠΈΠΉ ΠΊΠ»Π΅Ρ‚ΠΊΠΈ. ΠŸΡ€ΠΈΡ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Π½Ρ‹Π΅ Π±Π»ΠΎΠΊΠΈ ΡΠ°Ρ‚Π”ΠΠš Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌΡ‹ для ΠΏΡ€Π°Π²ΠΈΠ»ΡŒΠ½ΠΎΠΉ ассоциации сСстринских Ρ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ΄. ЦСлью настоящСй Ρ€Π°Π±ΠΎΡ‚Ρ‹ являлось ΠΈΠ·ΡƒΡ‡Π΅Π½ΠΈΠ΅ взаимодСйствия ассоциированных с ΡΠ°Ρ‚Π΅Π»Π»ΠΈΡ‚Π½Ρ‹ΠΌΠΈ Π”ΠΠš Π±Π΅Π»ΠΊΠΎΠ² SAF-A/hnRNP… Π§ΠΈΡ‚Π°Ρ‚ΡŒ Π΅Ρ‰Ρ‘ >

Π‘ΠΎΠ΄Π΅Ρ€ΠΆΠ°Π½ΠΈΠ΅

  • БПИБОК Π˜Π‘ΠŸΠžΠ›Π¬Π—Π£Π•ΠœΠ«Π₯ Π‘ΠžΠšΠ ΠΠ©Π•ΠΠ˜Π™
  • 1. Π’Π’Π•Π”Π•ΠΠ˜Π•
    • 1. 1. Π¦Π΅Π»ΠΈ ΠΈ Π·Π°Π΄Π°Ρ‡ΠΈ исслСдования
    • 1. 2. ΠžΡΠ½ΠΎΠ²Π½Ρ‹Π΅ полоТСния, выносимыС Π½Π° Π·Π°Ρ‰ΠΈΡ‚Ρƒ
  • 2. Π›Π˜Π’Π•Π ΠΠ’Π£Π ΠΠ«Π™ ΠžΠ‘Π—ΠžΠ 
    • 2. 1. ΠžΡ€Π³Π°Π½ΠΈΠ·Π°Ρ†ΠΈΡ Π³Π΅Π½ΠΎΠΌΠ° эукариот
    • 2. 2. ΠžΡΠΎΠ±Π΅Π½Π½ΠΎΡΡ‚ΠΈ ΠΎΡ€Π³Π°Π½ΠΈΠ·Π°Ρ†ΠΈΠΈ сатСллитных Π”ΠΠš
    • 2. 3. Π‘Π΅Π»ΠΊΠΈ, спСцифичСски ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΠ΅ ΡΠ°Ρ‚Π”ΠΠš
    • 2. 4. Π“Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½
  • 3. ΠœΠΠ’Π•Π Π˜ΠΠ›Π« И ΠœΠ•Π’ΠžΠ”Π«
    • 3. 1. БиохимичСскиС ΠΌΠ΅Ρ‚ΠΎΠ΄Ρ‹
      • 3. 1. 1. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ ΠΏΡ€Π΅ΠΏΠ°Ρ€Π°Ρ‚Π° ядСрного матрикса
      • 3. 1. 2. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ ядСрного экстракта ΠΈΠ· ΠΊΠ»Π΅Ρ‚ΠΎΠΊ ΠΊΡƒΠ»ΡŒΡ‚ΡƒΡ€Ρ‹ HeLa
      • 3. 1. 3. Аффинная очистка Π°ΠΊΡ‚ΠΈΠ½-ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΡ… Π±Π΅Π»ΠΊΠΎΠ² Π½Π°Π”ΠΠšΠ°Π·Π΅ I
      • 3. 1. 4. Аффинная очистка ядСрного Π°ΠΊΡ‚ΠΈΠ½Π°
      • 3. 1. 5. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ ΠΈ ΠΈΡΠΏΠΎΠ»ΡŒΠ·ΠΎΠ²Π°Π½ΠΈΠ΅ Ρ€Π°Π΄ΠΈΠΎΠ°ΠΊΡ‚ΠΈΠ²Π½ΠΎ ΠΌΠ΅Ρ‡Π΅Π½Π½Ρ‹Ρ… Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚ΠΎΠ² аминокислотной ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Π±Π΅Π»ΠΊΠ° SAF-A/hnRNP U
      • 3. 1. 6. Π˜Π·ΡƒΡ‡Π΅Π½ΠΈΠ΅ стСпСни аффиности ΠΏΠ΅ΠΏΡ‚ΠΈΠ΄ΠΎΠ² ΠΊ ΡΠ΄Π΅Ρ€Π½ΠΎΠΌΡƒ Π°ΠΊΡ‚ΠΈΠ½Ρƒ
      • 3. 1. 1. SDS-элСктрофорСз ΠΈ ΠΎΠΊΡ€Π°ΡˆΠΈΠ²Π°Π½ΠΈΠ΅ Π³Π΅Π»Π΅ΠΉ
    • 3. 2. ΠœΠΎΠ»Π΅ΠΊΡƒΠ»ΡΡ€Π½Ρ‹Π΅ ΠΌΠ΅Ρ‚ΠΎΠ΄Ρ‹
      • 3. 2. 1. ΠŸΠ»Π°Π·ΠΌΠΈΠ΄Ρ‹
      • 3. 2. 2. Π’Ρ‹Π΄Π΅Π»Π΅Π½ΠΈΠ΅ ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Π½ΠΎΠΉ Π”ΠΠš, элСктрофорСз Π”ΠΠš
      • 3. 2. 3. ΠŸΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·Π½Π°Ρ цСпная рСакция (ПЦР)
      • 3. 2. 4. ΠŸΡ€Π°ΠΉΠΌΠ΅Ρ€Ρ‹
      • 3. 2. 5. ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚ΠΎΠ² Π”ΠΠš
    • 3. 3. Π˜ΠΌΠΌΡƒΠ½ΠΎΠ»ΠΎΠ³ΠΈΡ‡Π΅ΡΠΊΠΈΠ΅ ΠΌΠ΅Ρ‚ΠΎΠ΄Ρ‹
  • 3. 3,1. АнтитСла
    • 3. 3. 2. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ ΠΏΠΎΠ»ΠΈΡ‚ΠΎΠ½Π°Π»ΡŒΠ½Ρ‹Ρ… Π°Π½Ρ‚ΠΈΡ‚Π΅Π»
    • 3. 3. 3. Π˜ΠΌΠΌΡƒΠ½ΠΎΠΏΡ€Π΅Ρ†ΠΈΠΏΠΈΡ‚Π°Ρ†ΠΈΡ
    • 3. 3. 4. Π˜ΠΌΠΌΡƒΠ½ΠΎΠ±Π»ΠΎΡ‚ΠΈΠ½Π³
    • 3. 4. ЦитологичСскиС ΠΌΠ΅Ρ‚ΠΎΠ΄Ρ‹
    • 3. 4. 1. ΠšΡƒΠ»ΡŒΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠ»Π΅Ρ‚ΠΎΠΊ
    • 3. 4. 2. ИспользованиС диотиобис-ΡΡƒΠΊΡ†ΠΈΠ½Π³ΡˆΠΈΠ΄ΠΈΠ»ΠΏΡ€ΠΎΠΏΠΈΠΎΠ½Π°Ρ‚Π°
    • 3. 4. 3. ВрансфСкция ΠΊΠ»Π΅Ρ‚ΠΎΠΊ ΠΊΡƒΠ»ΡŒΡ‚ΡƒΡ€Ρ‹
    • 3. 4. 4. Бинхронизация ΠΊΠ»Π΅Ρ‚ΠΎΠΊ
    • 3. 4. 5. Π˜ΠΌΠΌΡƒΠ½ΠΎΡ„Π»ΡƒΠΎΡ€Π΅ΡΡ†Π΅Π½Ρ†ΠΈΡ
    • 3. 4. 6. ΠœΠΈΠΊΡ€ΠΎΠΈΠ½ΡŠΠ΅ΠΊΡ†ΠΈΠΈ
    • 3. 4. 7. ВыявлСниС сайтов транскрипции
    • 3. 5. ΠœΠ΅Ρ‚ΠΎΠ΄Ρ‹ ΠΊΠΎΠΌΠΏΡŒΡŽΡ‚Π΅Ρ€Π½ΠΎΠ³ΠΎ Π°Π½Π°Π»ΠΈΠ·Π°
  • РЕЗУЛЬВАВЫ
    • 4. 1. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ ΠΈ Ρ‚СстированиС ΠΏΠΎΠ»ΠΈΠΊΠ»ΠΎΠ½Π°Π»ΡŒΠ½Ρ‹Ρ… Π°Π½Ρ‚ΠΈΡ‚Π΅Π» ΠΏΡ€ΠΎΡ‚ΠΈΠ² Π±Π΅Π»ΠΊΠ° NAP5 7/D yskerin
    • 4. 2. Локализация Π±Π΅Π»ΠΊΠΎΠ² Π¨Π Π 140 ΠΈ NAP57/Dyskerin
    • 4. 3. ВнутриядСрная локализация Π±Π΅Π»ΠΊΠ° NAP57/Dyskerin
    • 4. 4. Локализация Π±Π΅Π»ΠΊΠ° NAP57 ΠΈ Π ΠΠš-Π³Π΅Π»ΠΈΠΊΠ°Π·Ρ‹ Ρ€68 сСмСйства DEAD
    • 4. 5. Π”ΠΎΠΌΠ΅Π½Π½Ρ‹ΠΉ Π°Π½Π°Π»ΠΈΠ· ΠΏΠ΅Ρ€Π²ΠΈΡ‡Π½ΠΎΠΉ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Π±Π΅Π»ΠΊΠ° SAF-A/hnRNPU
    • 4. 6. Π˜Π·ΡƒΡ‡Π΅Π½ΠΈΠ΅ Ρ„ΡƒΠ½ΠΊΡ†ΠΈΠΎΠ½Π°Π»ΡŒΠ½ΠΎΠΉ активности ΠΈ Ρ…Π°Ρ€Π°ΠΊΡ‚Π΅Ρ€Π° Π»ΠΎΠΊΠ°Π»ΠΈΠ·Π°Ρ†ΠΈΠΈ ΠΎΡ‚Π΄Π΅Π»ΡŒΠ½Ρ‹Ρ… Π΄ΠΎΠΌΠ΅Π½ΠΎΠ² Π±Π΅Π»ΠΊΠ° SAF-A, ΠΌΠ΅Ρ‡Π΅Π½Π½Ρ‹Ρ… GFP
    • 4. 7. Π‘Π΅Π»ΠΎΠΊ SAF-A взаимодСйствуСт с ΡΠ΄Π΅Ρ€Π½Ρ‹ΠΌ Π -Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ in vitro ΠΈ in vivo
    • 4. 8. Локализация Π±Π΅Π»ΠΊΠ° SAF-A ΠΈ ΡΠ΄Π΅Ρ€Π½ΠΎΠ³ΠΎ Π°ΠΊΡ‚ΠΈΠ½Π° Π² ΠΈΠ½Ρ‚Π΅Ρ€Ρ„Π°Π·Π½Ρ‹Ρ… ΠΈ ΠΌΠΈΡ‚отичСских ΠΊΠ»Π΅Ρ‚ΠΊΠ°Ρ…
    • 4. 9. Π‘Π΅Π»ΠΎΠΊ SAF-A взаимодСйствуСт с Π -Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Π² ΡΠ΄Ρ€Π΅ Ρ‡Π΅Ρ€Π΅Π· Π‘-ΠΊΠΎΠ½Ρ†Π΅Π²ΠΎΠΉ Π΄ΠΎΠΌΠ΅Π½ Π±Π΅Π»ΠΊΠ°
    • 4. 10. Π‘Π΅Π»ΠΎΠΊ SAF-A взаимодСйствуСт с Π -Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Π² ΡΠ΄Ρ€Π΅ Ρ‡Π΅Ρ€Π΅Π· пяти Π°ΠΌΠΈΠ½ΠΎΠΊΠΈΡΠ»ΠΎΡ‚Π½ΡƒΡŽ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΡŒ QRTQK Π½Π° Π‘-ΠΊΠΎΠ½Ρ†Π΅
    • 4. 11. КомплСкс SAF-A с Π -Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌ для транскрипции РНК-ΠΏΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·ΠΎΠΉII
  • 5. ΠžΠ‘Π‘Π£Π–Π”Π•ΠΠ˜Π• Π Π•Π—Π£Π›Π¬Π’ΠΠ’ΠžΠ’
  • Π’Π«Π’ΠžΠ”Π«

ВзаимодСйствиС ядСрного Π°ΠΊΡ‚ΠΈΠ½Π° ΠΈ NAP57/дискСрина с Π±Π΅Π»ΠΊΠ°ΠΌΠΈ, спСцифичСски ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΠΌΠΈ ΡΠ°Ρ‚Π΅Π»Π»ΠΈΡ‚Π½ΡƒΡŽ Π”ΠΠš (Ρ€Π΅Ρ„Π΅Ρ€Π°Ρ‚, курсовая, Π΄ΠΈΠΏΠ»ΠΎΠΌ, ΠΊΠΎΠ½Ρ‚Ρ€ΠΎΠ»ΡŒΠ½Π°Ρ)

Π‘ΠΎ ΡΡ‚Ρ€ΡƒΠΊΡ‚ΡƒΡ€Π½ΠΎΠΉ Ρ‚ΠΎΡ‡ΠΊΠΈ зрСния, классичСский конститутивный Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½ прСдставляСт собой ΠΏΠΎΠ²Ρ‚ΠΎΡ€ΡΡŽΡ‰ΠΈΠ΅ΡΡ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Ρ‚Π°Π½Π΄Π΅ΠΌΠ½ΠΎ располоТСнных ΠΌΠΎΠ½ΠΎΠΌΠ΅Ρ€ΠΎΠ² сатСллитных Π”ΠΠš (ΡΠ°Ρ‚Π”ΠΠš), ассоциированныС со ΡΠΏΠ΅Ρ†ΠΈΡ„ичСскими Π±Π΅Π»ΠΊΠ°ΠΌΠΈ. Π£ Π²Ρ‹ΡΡˆΠΈΡ… эукариот ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ ΡΠ°Ρ‚Π”ΠΠš ΠΊΠΎΠ½Ρ†Π΅Π½Ρ‚Ρ€ΠΈΡ€ΡƒΡŽΡ‚ΡΡ Π² Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Π½Ρ‹Ρ… ΠΈ Ρ‚Π΅Π»ΠΎΠΌΠ΅Ρ€Π½Ρ‹Ρ… Ρ€Π°ΠΉΠΎΠ½Π°Ρ… хромосом, ΠΎΠ΄Π½Π°ΠΊΠΎ ΠΈΡ… Ρ„ΡƒΠ½ΠΊΡ†ΠΈΠΎΠ½Π°Π»ΡŒΠ½Π°Ρ Ρ€ΠΎΠ»ΡŒ Π² ΠΏΡ€ΠΎΡ†Π΅ΡΡΠ΅ Ρ€Π΅Π°Π»ΠΈΠ·Π°Ρ†ΠΈΠΈ гСнСтичСской ΠΈΠ½Ρ„ΠΎΡ€ΠΌΠ°Ρ†ΠΈΠΈ Π΄ΠΎ ΡΠΈΡ… ΠΏΠΎΡ€ Π½Π΅ ΡΡΠ½Π°.

ДостовСрно извСстно, Ρ‡Ρ‚ΠΎ ΡΠ°Ρ‚Π”ΠΠš, ΠΎΠ±Ρ€Π°Π·ΡƒΡŽΡ‰ΠΈΠ΅ Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Ρƒ, ΠΎΠ±Π΅ΡΠΏΠ΅Ρ‡ΠΈΠ²Π°ΡŽΡ‚ Ρ„ΠΎΡ€ΠΌΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΈΠ½Π΅Ρ‚ΠΎΡ…ΠΎΡ€Π°, ΠΊΠΎΡ€Ρ€Π΅ΠΊΡ‚Π½ΠΎΠ΅ спариваниС ΠΈ Ρ€Π°Π·Π΄Π΅Π»Π΅Π½ΠΈΠ΅ хромосом Π² Ρ…ΠΎΠ΄Π΅ митотичСского ΠΈ ΠΌΠ΅ΠΉΠΎΡ‚ичСского Π΄Π΅Π»Π΅Π½ΠΈΠΉ ΠΊΠ»Π΅Ρ‚ΠΊΠΈ. ΠŸΡ€ΠΈΡ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Π½Ρ‹Π΅ Π±Π»ΠΎΠΊΠΈ ΡΠ°Ρ‚Π”ΠΠš Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌΡ‹ для ΠΏΡ€Π°Π²ΠΈΠ»ΡŒΠ½ΠΎΠΉ ассоциации сСстринских Ρ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ΄.

Π’ ΠΈΠ½Ρ‚Π΅Ρ€Ρ„Π°Π·Π½Ρ‹ΠΉ ΠΏΠ΅Ρ€ΠΈΠΎΠ΄ ΡΠ°Ρ‚Π”ΠΠš участвуСт Π² Ρ‚Ρ€Π΅Ρ…ΠΌΠ΅Ρ€Π½ΠΎΠΉ ΠΎΡ€Π³Π°Π½ΠΈΠ·Π°Ρ†ΠΈΠΈ ΠΈΠ½Ρ‚Π΅Ρ€Ρ„Π°Π·Π½ΠΎΠ³ΠΎ ядра (Razin et al., 1995; Manuelidis, 1997), ΠΎΠΏΡ€Π΅Π΄Π΅Π»ΡΡŽΡ‰ΡƒΡŽ транскрипционный ΠΏΠ°Ρ‚Ρ‚Π΅Ρ€Π½ ΠΊΠ»Π΅Ρ‚ΠΊΠΈ. Π˜Π·Π²Π΅ΡΡ‚Π½ΠΎ, Ρ‡Ρ‚ΠΎ Ρ…Π°Ρ€Π°ΠΊΡ‚Π΅Ρ€ распрСдСлСния Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€ Π² ΡΠ΄Ρ€Π΅ мСняСтся Π² Ρ…ΠΎΠ΄Π΅ ΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½ΠΎΠ³ΠΎ Ρ†ΠΈΠΊΠ»Π° (Ferguson, Ward, 1992) ΠΈ ΡΠΌΠ±Ρ€ΠΈΠΎΠ½Π°Π»ΡŒΠ½ΠΎΠ³ΠΎ развития (Longo et al., 2003). ИзмСнСниС полоТСния Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€ Π² ΡΠ΄Ρ€Π°Ρ… Π½Π΅ΠΉΡ€ΠΎΠ½ΠΎΠ² происходит ΠΏΡ€ΠΈ ΠΈΠ·ΠΌΠ΅Π½Π΅Π½ΠΈΠΈ транскрипционной активности ΠΏΡ€ΠΈ ростС аксонов Π² ΡƒΡΠ»ΠΎΠ²ΠΈΡΡ… in vitro (Chon, De Boni, 1996). Π₯ΠΎΡ€ΠΎΡˆΠΎ извСстна ΡΠΏΠΎΡΠΎΠ±Π½ΠΎΡΡ‚ΡŒ Ρ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Π½ΠΎΠ³ΠΎ Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½Π° ΠΈΠ½Π³ΠΈΠ±ΠΈΡ€ΠΎΠ²Π°Ρ‚ΡŒ Ρ‚Ρ€Π°Π½ΡΠΊΡ€ΠΈΠΏΡ†ΠΈΠΎΠ½Π½ΡƒΡŽ Π°ΠΊΡ‚ΠΈΠ²Π½ΠΎΡΡ‚ΡŒ Π³Π΅Π½ΠΎΠ², ΠΏΡ€ΠΈ ассоциации послСдних с Ρ€Π°ΠΉΠΎΠ½Π°ΠΌΠΈ Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½Π°. Π‘ Π΄Ρ€ΡƒΠ³ΠΎΠΉ стороны, ΠΈΠ½Π³ΠΈΠ±ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ РНК-ΠΏΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·Ρ‹ II ΠΏΡ€ΠΈΠ²ΠΎΠ΄ΠΈΡ‚ ΠΊ Π΄Π΅ΠΊΠΎΠ½Π΄Π΅Π½ΡΠ°Ρ†ΠΈΠΈ ΠΏΡ€ΠΈΡ†Π΅Π½Ρ‚Ρ€ΠΎΠΌΠ΅Ρ€Π½ΠΎΠ³ΠΎ Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½Π° (Haaf, Ward, 1996). НСдавниС исслСдования ΠΏΠΎΠΊΠ°Π·Π°Π»ΠΈ ΠΏΡ€ΡΠΌΡƒΡŽ связь ΠΌΠ΅ΠΆΠ΄Ρƒ Ρ„ΠΎΡ€ΠΌΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ΠΌ Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½Π° ΠΈ ΠΏΠΎΡΡ‚гранскрипционной рСгуляциСй Π³Π΅Π½Π½ΠΎΠΉ активности посрСдством ΠΌΠ΅Ρ…Π°Π½ΠΈΠ·ΠΌΠ° RNA interference (RNAi) (Agrawal et al., 2003), подтвСрТдая участиС Π³Π΅Ρ‚Π΅Ρ€ΠΎΡ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½ΠΎΠ²Ρ‹Ρ… Ρ€Π°ΠΉΠΎΠ½ΠΎΠ² хромосом Π² ΠΌΠ΅Ρ‚Π°Π±ΠΎΠ»ΠΈΠ·ΠΌΠ΅ ΠΌΠΎΠ»Π΅ΠΊΡƒΠ» РНК.

ΠΠΊΡ‚ΠΈΠ²Π½ΠΎΡΡ‚ΡŒ ΡΠ°Ρ‚Π”ΠΠš Π΄ΠΎΠ»ΠΆΠ½Π° ΠΎΠΏΡ€Π΅Π΄Π΅Π»ΡΡ‚ΡŒΡΡ Π±Π΅Π»ΠΊΠΎΠ²Ρ‹ΠΌΠΈ Ρ„Π°ΠΊΡ‚ΠΎΡ€Π°ΠΌΠΈ, способными спСцифичСски Ρ€Π°ΡΠΏΠΎΠ·Π½Π°Π²Π°Ρ‚ΡŒ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ ΡΠ°Ρ‚Π”ΠΠš Π² ΡΠ΄Ρ€Π΅ ΠΈΠ½Ρ‚Π΅Ρ€Ρ„Π°Π·Π½ΠΎΠΉ ΠΊΠ»Π΅Ρ‚ΠΊΠΈ. ΠœΠ΅ΠΆΠ΄Ρƒ Ρ‚Π΅ΠΌ, Π±Π΅Π»ΠΊΠΈ ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΠ΅ ΡΠ°Ρ‚Π”ΠΠš ΠΈΠ·ΡƒΡ‡Π΅Π½Ρ‹ нСдостаточно ΠΏΠΎΠ»Π½ΠΎ. Π˜Π·Π²Π΅ΡΡ‚Π½Ρ‹Π΅ ΡΠ°Ρ‚Π”ΠΠš-ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΠ΅ Π±Π΅Π»ΠΊΠΈ ΠΎΠΏΡ€Π΅Π΄Π΅Π»Π΅Π½Ρ‹ с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ сыворотки Π°ΡƒΡ‚ΠΎΠΈΠΌΠΌΡƒΠ½Π½Ρ‹Ρ… Π±ΠΎΠ»ΡŒΠ½Ρ‹Ρ… (Moroi et al., 1980; Earnshaw, Rothfield, 1985), Π»ΠΈΠ±ΠΎ ΠΌΠ΅Ρ‚ΠΎΠ΄ΠΎΠΌ Ρ€Π΅Ρ‚Π°Ρ€Π΄Π°Ρ†ΠΈΠΈ Π² Π³Π΅Π»Π΅ (Lobov et al., 1998), основанным Π½Π° ΠΈΠ·ΠΌΠ΅Π½Π΅Π½ΠΈΠΈ подвиТности Π”ΠΠš-Π±Π΅Π»ΠΊΠΎΠ²ΠΎΠ³ΠΎ комплСкса Π² ΠŸΠΠΠ“.

ПослСдний ΠΌΠ΅Ρ‚ΠΎΠ΄ Π½Π°ΠΈΠ±ΠΎΠ»Π΅Π΅ эффСктивСн, ΠΏΠΎΡΠΊΠΎΠ»ΡŒΠΊΡƒ позволяСт Ρ‚ΠΎΡ‡Π½ΠΎ ΠΎΠΏΡ€Π΅Π΄Π΅Π»ΠΈΡ‚ΡŒ ΡΠΏΠ΅Ρ†ΠΈΡ„ΠΈΡ‡Π½ΠΎΡΡ‚ΡŒ распознавания ΠΈΡΠΏΠΎΠ»ΡŒΠ·ΠΎΠ²Π°Π½Π½Ρ‹Ρ… Π² ΠΊΠ°Ρ‡Π΅ΡΡ‚Π²Π΅ Π·ΠΎΠ½Π΄Π° ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚Π΅ΠΉ Π”ΠΠš. Π’Π°ΠΊ с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ ΠΌΠ΅Ρ‚ΠΎΠ΄Π° Ρ€Π΅Ρ‚Π°Ρ€Π΄Π°Ρ†ΠΈΠΈ Π² Π³Π΅Π»Π΅ Π±Ρ‹Π» установлСн спСцифичСский Ρ…Π°Ρ€Π°ΠΊΡ‚Π΅Ρ€ взаимодСйствия РНК-Π³Π΅Π»ΠΈΠΊΠ°Π·Ρ‹ Ρ€68 сСмСйства DEAD с ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΡΠΌΠΈ Π°Π»ΡŒΡ„ΠΎΠΈΠ΄Π½ΠΎΠΉ Π”ΠΠš (Π°-ΡΠ°Ρ‚Π”ΠΠš) Ρ‡Π΅Π»ΠΎΠ²Π΅ΠΊΠ° ΠΈ ΠΌΠΈΠ½ΠΎΡ€Π½ΠΎΠ³ΠΎ сатСллита (ΠΌΠΈΠ‘Π°Ρ‚) ΠΌΡ‹ΡˆΠΈ (Podgornaya et al., 2003; Enukashvily et al., 2005), Π° Ρ‚Π°ΠΊΠΆΠ΅ взаимодСйствия ядСрного Π±Π΅Π»ΠΊΠ° SAF-A/hnRNP U Ρ ΠΌΠ°ΠΆΠΎΡ€Π½Ρ‹ΠΌ сатСллитом ΠΌΡ‹ΡˆΠΈ (ΠΌΠ°Π‘Π°Ρ‚) (Lobov et al., 2000; 2001).

ΠŸΡ€Π΅Π΄ΠΌΠ΅Ρ‚ΠΎΠΌ настоящСго исслСдования стало ΠΈΠ·ΡƒΡ‡Π΅Π½ΠΈΠ΅ Π±Π΅Π»ΠΊΠΎΠ²Ρ‹Ρ… комплСксов, Π² ΡΠΎΡΡ‚Π°Π² ΠΊΠΎΡ‚ΠΎΡ€Ρ‹Ρ… входят РНК-Π³Π΅Π»ΠΈΠΊΠ°Π·Π° Ρ€68 ΠΈ Π±Π΅Π»ΠΎΠΊ SAF-A/hnRNP U.

1.1. Π¦Π΅Π»ΠΈ ΠΈ Π·Π°Π΄Π°Ρ‡ΠΈ исслСдования.

ЦСлью настоящСй Ρ€Π°Π±ΠΎΡ‚Ρ‹ являлось ΠΈΠ·ΡƒΡ‡Π΅Π½ΠΈΠ΅ взаимодСйствия ассоциированных с ΡΠ°Ρ‚Π΅Π»Π»ΠΈΡ‚Π½Ρ‹ΠΌΠΈ Π”ΠΠš Π±Π΅Π»ΠΊΠΎΠ² SAF-A/hnRNP U ΠΈ Π ΠΠš-Π³Π΅Π»ΠΈΠΊΠ°Π·Ρ‹ Ρ€68 с Π½Π΅Ρ…Ρ€ΠΎΠΌΠ°Ρ‚ΠΈΠ½ΠΎΠ²Ρ‹ΠΌΠΈ Π±Π΅Π»ΠΊΠ°ΠΌΠΈ — ядСрным fJ-Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ ΠΈ Π±Π΅Π»ΠΊΠΎΠΌ N ΠΠ 5 7/Dyskerin.

Π‘Ρ‹Π»ΠΈ поставлСны ΡΠ»Π΅Π΄ΡƒΡŽΡ‰ΠΈΠ΅ Π·Π°Π΄Π°Ρ‡ΠΈ:

1. ΠŸΠΎΠ»ΡƒΡ‡ΠΈΡ‚ΡŒ Π°Π½Ρ‚ΠΈΡ‚Π΅Π»Π°, спСцифичСски ΡƒΠ·Π½Π°ΡŽΡ‰ΠΈΠ΅ Π±Π΅Π»ΠΎΠΊ NAP57.

2. Π˜Π·ΡƒΡ‡ΠΈΡ‚ΡŒ Π²Π½ΡƒΡ‚Ρ€ΠΈΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½ΡƒΡŽ Π»ΠΎΠΊΠ°Π»ΠΈΠ·Π°Ρ†ΠΈΡŽ ΡΠ°Ρ‚Π”ΠΠš-ΡΠ²ΡΠ·Ρ‹Π²Π°ΡŽΡ‰ΠΈΡ… ΠΈ ΡΠΎΠΏΡƒΡ‚ΡΡ‚Π²ΡƒΡŽΡ‰ΠΈΡ… ΠΈΠΌ Π±Π΅Π»ΠΊΠΎΠ².

3. ΠŸΠΎΠ΄Ρ‚Π²Π΅Ρ€Π΄ΠΈΡ‚ΡŒ взаимодСйствиС ΠΏΠ°Ρ€ молСкулярных ΠΏΠ°Ρ€Ρ‚Π½Π΅Ρ€ΠΎΠ² (Π³Π΅Π»ΠΈΠΊΠ°Π·Π° p68/NAP-57 ΠΈ SAF-A/p-Π°ΠΊΡ‚ΠΈΠ½) in vivo ΠΈ in vitro.

4. Π˜Π΄Π΅Π½Ρ‚ΠΈΡ„ΠΈΡ†ΠΈΡ€ΠΎΠ²Π°Ρ‚ΡŒ Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌΡ‹Π΅ для формирования Π±Π΅Π»ΠΊΠΎΠ²Ρ‹Ρ… комплСксов аминокислотныС ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ.

5. ΠžΠΏΡ€Π΅Π΄Π΅Π»ΠΈΡ‚ΡŒ Ρ„ΡƒΠ½ΠΊΡ†ΠΈΠΈ ΠΈΠ·ΡƒΡ‡Π°Π΅ΠΌΡ‹Ρ… Π±Π΅Π»ΠΊΠΎΠ²Ρ‹Ρ… комплСксов.

Π²Ρ‹Π²ΠΎΠ΄Ρ‹.

1. ВнутриядСрная локализация Π±Π΅Π»ΠΊΠ° NAP57/Dyskerin Π½Π΅ Π·Π°Π²ΠΈΡΠΈΡ‚ ΠΎΡ‚ Ρ„Π°Π· ΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½ΠΎΠ³ΠΎ Ρ†ΠΈΠΊΠ»Π° ΠΈ ΠΌΠΎΠΆΠ΅Ρ‚ Π±Ρ‹Ρ‚ΡŒ Ρ€Π°Π·Π΄Π΅Π»Π΅Π½Π° Π½Π° Π΄Π²Π° Ρ‚ΠΈΠΏΠ°: ΡΠ΄Ρ€Ρ‹ΡˆΠΊΠΎΠ²Π°Ρ ΠΈ Π½ΡƒΠΊΠ»Π΅ΠΎΠΏΠ»Π°Π·ΠΌΠ΅Π½Π½Π°Ρ.

2. Π‘Π΅Π»ΠΎΠΊ NAP57/Dyskerin спСцифичСски взаимодСйствуСт in vitro с Π ΠΠš-Π³Π΅Π»ΠΈΠΊΠ°Π·ΠΎΠΉ Ρ€68 сСмСйства DEAD, ΠΊΠΎΠ»ΠΎΠΊΠ°Π»ΠΈΠ·ΡƒΡΡΡŒ с Π½Π΅ΠΉ Π²ΠΎ Π²Π½ΡƒΡ‚риядСрных Ρ…Ρ€Π°Π½ΠΈΠ»ΠΈΡ‰Π°Ρ… Ρ„Π°ΠΊΡ‚ΠΎΡ€ΠΎΠ² сплайсинга (спСклах) in vivo.

3. Π‘Π΅Π»ΠΎΠΊ SAF-A спСцифичСски взаимодСйствуСт с (3-Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Π² ΡΠ΄Ρ€Π΅, образуя Π΅Π΄ΠΈΠ½Ρ‹ΠΉ комплСкс с Π ΠΠš-ΠΏΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·ΠΎΠΉ II.

4. Π‘Π΅Π»ΠΎΠΊ SAF-A взаимодСйствуСт с ΡΠ΄Π΅Ρ€Π½Ρ‹ΠΌ (3-Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Ρ‡Π΅Ρ€Π΅Π· ΠΏΡΡ‚ΠΈΠ°ΠΌΠΈΠ½ΠΎΠΊΠΈΡΠ»ΠΎΡ‚Π½ΡƒΡŽ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΡŒ: Π³Π»ΡƒΡ‚Π°ΠΌΠΈΠ½-Π°Ρ€Π³ΠΈΠ½ΠΈΠ½-Ρ‚Ρ€Π΅ΠΎΠ½ΠΈΠ½-Π³Π»ΡƒΡ‚Π°ΠΌΠΈΠ½-Π»ΠΈΠ·ΠΈΠ½ Π½Π° Π‘-ΠΊΠΎΠ½Ρ†Π΅ аминокислотной ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ SAF-А.

5. КомплСкс SAF-A с Ρ€-Π°ΠΊΡ‚ΠΈΠ½ΠΎΠΌ Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌ для транскрипции РНК-ΠΏΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·ΠΎΠΉ II.

ΠŸΠΎΠΊΠ°Π·Π°Ρ‚ΡŒ вСсь тСкст

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