Sains Malaysiana 51(11)(2022): 3621-3633

http://doi.org/10.17576/jsm-2022-5111-09

 

Expression Analysis using Reverse Transcription Quantitative Real-Time PCR (RT-qPCR) Suggests Different Strategies in Parageobacillus caldoxylosilyticus ER4B under Exposure to Cold Shock

(Analisis Pengekspresan menggunakan Transkripsi Berbalik Kuantitatif Masa-Nyata PCR (RT-qPCR) Mencadangkan Strategi Berbeza untuk Parageobacillus caldoxylosilyticus ER4B di bawah Pendedahan kepada Renjatan Sejuk)

 

CHING XIN JIE1, NOOR HYDAYATY MD YUSUF1, NAZALAN NAJIMUDIN 2, CHEAH YOKE KQUEEN3 & CLEMENTE MICHAEL WONG VUI LING1,4,*

 

1Biotechnology Research Institute, Universiti Malaysia Sabah, 88400 Kota Kinabalu, Sabah, Malaysia

2School of Biological Science, Universiti Sains Malaysia, Persiaran Bukit Jambul, 11900 Bayan Lepas, Penang, Malaysia

3Department of Biomedical Science, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia

4National Antarctic Research Centre, Universiti Malaya, 50603 Kuala Lumpur, Federal Territory, Malaysia

 

Diserahkan: 29 April 2021/Diterima: 5 Julai 2022

 

Abstract

Microorganisms have acquired both common and unique abilities to withstand cold stress on Earth. Many studies on bacterial cold shock have been conducted, however, the majority of the studies were focused on mesophiles and psychrophiles. To date, limited information is available on the response of thermophilic bacteria to cold stress and therefore, it is not known how thermophilic bacteria would respond to different cold shocks. To address this question, the cold shock responses of a thermophilic Parageobacillus caldoxylosilyticus ER4B which has an optimal growth temperature at 64 °C were determined using Real-Time PCR (RT-qPCR). When the bacterium was exposed to mild cold shock at 54 °C, the expressions of gene encoding for pyruvate kinase and acetolactate synthase were significantly upregulated, suggesting that more pyruvate molecules were produced to synthesize branched-chain amino acids that could alter the fatty acid profile on the cell membrane. Accumulation of pyruvate in the bacterium could also help to scavenge cold-induced reactive oxygen species (ROS). Meanwhile, exposing the bacterium to extreme cold shock at 10 °C resulted in significant upregulation of genes encoding for γ-glutamylcyclotransferase, cold shock protein B and competence protein ComEA. An increase in these enzymes expression indicated more extreme measures including apoptosis and transformation were adopted during extreme cold shock.

 

Keywords: Cold shock; cold stress response; Parageobacillus caldoxylosilyticus; reverse transcription quantitative real-time PCR (RT-qPCR); thermophilic bacterium

 

Abstrak

Mikroorganisma telah memperoleh ciri umum dan unik untuk bertahan daripada tekanan suhu sejuk di bumi. Banyak kajian mengenai kejutan sejuk bakteria telah dijalankan, namun kebanyakannya memfokuskan kepada bakteria mesofil dan psikofil. Sehingga kini, terdapat maklumat yang terhad mengenai gerak balas bakteria termofilik terhadap tekanan suhu rendah dan sehubungan itu, bagaimana bakteria termofilik bergerak balas terhadap kejutan sejuk yang berbeza masih belum diketahui. Untuk menjawab persoalan ini, gerak balas kejutan sejuk daripada termofilik Parageobacillus caldoxylosilyticus ER4B yang mempunyai suhu pertumbuhan optimum pada suhu 64 °C telah ditentukan melalui PCR Masa-Nyata (RT-qPCR). Apabila bakteria terdedah kepada kejutan sejuk 54 °C, pengekspresan gen yang mengekodkan piruvat kinase dan asetolaktat sintase menunjukkan peningkatan transkrip, mencadangkan bahawa lebih banyak molekul piruvat dihasilkan untuk mensintesis asid amino rantai bercabang yang dapat mengubah profil asid lemak pada membran sel. Pengumpulan piruvat pada bakteria juga dapat membantu menghapuskan spesies oksigen reaktif (ROS) yang disebabkan oleh suhu rendah. Sementara itu, pendedahan bakteria kepada kejutan sejuk yang melampau pada 10 °C mengakibatkan peningkatan pengawalaturan pengekodan gen untuk γ-glutamil siklotransferase, protein kejutan sejuk B dan protein kompetensi ComEA. Peningkatan pengekspresan enzim tersebut menunjukkan pendekatan yang lebih ekstrem termasuk apoptosis dan transformasi dilakukan semasa kejutan sejuk yang melampau.

 

Kata kunci: Bakteria termofilik; gerak balas tekanan sejuk; kejutan sejuk; Parageobacillus caldoxylosilyticus; PCR masa nyata-kuantitatif transkripsi berbalik

 

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*Pengarang untuk surat-menyurat; email: michaelw@ums.edu.my

 

 

 

 

   

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