Background
Every organism faces changing environmental conditions and has evolved cellular machinery for coping with stress and adapting to changing environments. Change in temperature is one of the most common stresses faced by all living organisms. To respond to harmful effects of temperature downshift, there exists a family of proteins called cold shock proteins that play a significant role in acclimation of cells to cold [
1‐
5]. They help the cells to adapt and have pleiotropic functions inside the cell [
6,
7]. Cold shock proteins are among the most evolutionarily conserved proteins and are characterized by the presence of one or more cold shock domains (CSDs). CSDs have nucleic acid binding properties that bestow these proteins with several functions, including regulation of transcription, translation and splicing [
5,
8].
At low temperatures, cold shock proteins function as RNA chaperones by destabilizing secondary structures in target RNA. This enables the maintenance of single stranded state of target RNA to pursue efficient transcription and translation [
4,
9]. Cold shock proteins prevent formation of hairpin structures in RNA and, therefore, act as transcription anti-terminators [
10,
11].
Identification of cold shock proteins in bacteria
Cold shock proteins were initially found when a sudden drop in temperature (from 37 °C to 10 °C) caused a many-fold increase in the expression of a cold shock protein A (CspA) in
Escherichia coli [
12,
13]. Thereafter, cold shock proteins have been identified in several bacteria, including psychrophilic, mesophilic, thermophilic, and even hyperthermophilic bacteria [
4,
13,
14]. Sequence analysis indicates that bacterial cold shock proteins are small proteins with a molecular mass of approximately 7.4 kDa [
15] and comprise a typical CSD. They all have the ability to bind single-stranded RNA and DNA but no double-stranded DNA [
9,
16‐
19]. This protein-nucleic acid interaction is mediated by the moderately well-conserved nucleic acid binding motifs RNP1 (K/R-G-F/Y-G/A-F-V/I-X-F/Y) and RNP2 (L/I-F/Y-V/I-G/K-N/G-L) [
2,
20‐
22].
In
Escherichia coli, several cold induced proteins are expressed that include these cold shock proteins apart from RNA helicase csdA [
23], exoribonucleases PNPase and RNase R [
24], initiation factors 2a and 2b, NusA and RecA [
25]. Later, it was found that
Escherichia coli encodes 9 cold shock protein genes (
CspA to C
spI) that share 46–91% amino acid sequence similarity [
1]. Naming of cold shock proteins is done in similar fashion to other bacteria, however identical names do not necessarily share identical function and structure in different bacteria. Among all cold shock proteins,
CspC is constitutively expressed [
26] whereas
CspA,
CspB,
CspE, CspG, and
CspI are induced by cold shock [
27‐
31]. In contrast
CspD is induced by stationary phase growth and nutrient starvation [
32,
33] and
cspF and
cspH expression are not linked with any particular growth condition and their functions are not known [
34].
CspC and
CspE are known to regulate the expression of stress response proteins ‘RpoS’ and ‘UspA’ [
35] while
CspD is implicated in persister cell formation, biofilm development and inhibits DNA replication [
36,
37].
CspA is the major cold shock protein and the most prominent one in
Escherichia coli [
38]. A report by Giuliodori et al. suggested that mRNA of
CspA adopts different structures at low temperature which makes it less prone to degradation [
39]. As a result
CspA mRNA is translated more efficiently upon temperature fluctuation than
CspA mRNA at 37 °C [
39]. Likewise,
ttcsp2 of thermophilic
Thermus thermophilus was also reported to be cold induced protein that adopt more stable secondary structure in response to temperature drop [
40]. At 37 °C, the
cspA mRNA is very unstable, and has a half-life of only 12 s. Upon cold stress, its stability is dramatically increased as its half-life is now more than 20 min [
41]. This transient stabilization of
cspA mRNA on temperature drop implicates its significance in induction during cold shock [
42].
Xia et al. suggested that the functions of the CspA family members overlap and can compensate for each other [
31]. The authors found that by deleting 4 cold shock protein genes (
cspA, cspB, cspE, cspG) in
E. coli, a cold-sensitive strain ‘BX04’ was obtained that was unable to form colonies at 15 °C [
31]. The cold sensitivity of this strain can be suppressed by overexpressing any of the
E. coli cold shock protein genes, except
cspD. Moreover, loss of one or two cold shock protein genes in
E. coli increased the production of the remaining cold-induced cold shock protein genes. Similarly, in
Bacillus subtilis, deletion of one or two cold shock protein genes boosted the expression of remaining cold shock proteins post-cold shock [
43].
The three-dimensional structures of several bacterial cold shock proteins have been determined [
44‐
47]. Some of these include CspA from the mesophilic bacterium
Escherichia coli (EcCspA), cold shock protein from the thermophilic bacterium
Bacillus caldolyticus (BcCsp) and cold shock protein from the hyperthermophilic bacterium
Thermotoga maritima (TmCsp) [
44‐
47]. Structural studies indicate that cold shock proteins belong to oligonucleotide/oligosaccharide-binding (OB)-fold family of proteins. OB-fold consists of 5 antiparallel beta strands that form a Greek-key beta-barrel. Knowledge of OB-folded nucleoprotein complexes was found to originate from the X-ray structures of telomere DNA-binding proteins [
48‐
50]. Although structurally cold shock proteins are conserved, their thermo-stability differs [
14,
51]. Cold shock protein of thermophilic
Thermus aquaticus has a melting temperature of 76 °C and a rigid structure. On the contrary, CspA of the psychrotrophic
Listeria monocytogenes has a melting temperature of 40 °C [
51]. This implicates that psychrophilic cold shock proteins require higher structural flexibility to bind nucleic acids upon cold shock [
14].
Cold shock proteins in humans
The human genome encodes for about 8 members of cold shock genes namely YBX1, YBX2, YBX3, CARHSP1, CSDC2, CSDE1, LIN28A, and LIN28B [
52]. The best characterized members are denoted Y-box binding protein family. The prototypic member is Y-box binding protein-1 (YB-1), encoded by the gene YBX1. Two other members of Y-box binding protein family exist namely DNA binding protein A (DbpA) and C (DbpC) that are encoded by the genes YBX3 and YBX2, respectively [
52]. YB-1 was first named in 1988 to refer to transcription factors that interact with the Ybox motif in the promoter of the major histocompatibility complex class II genes [
53]. YB-1 has properties of a nucleic acid chaperone and binds with both DNA and RNA. By its nuclei acid binding ability, it is involved in several mRNA- and DNA-dependent processes, including mRNA splicing, mRNA translation, DNA replication and repair [
54‐
59]. The protein functions as a positive transcription factor to upregulate several genes, including MDR1 (multi-drug resistance-1) [
58]. The MDR1 promoter activity is known to increase in response to various environmental stimuli, including anticancer agents and ultraviolet irradiation [
60,
61]. YB-1 also increases resistance of cells to ionizing radiation and xenobiotics when involved in DNA repair in the nucleus [
61,
62]. YB-1 nuclear localization is therefore considered an early marker of multidrug resistance of malignant cells [
62‐
64].
Another important cold shock protein expressed in humans is calcium-regulated heat-stable protein 1 (CARHSP1); a 24 kDa protein also known as CRHSP-24. CARHSP1 binds to tumor necrosis factor (TNF) mRNA and play a role in its stabilization within P-bodies and exosomes [
65]. It is dephosphorylated by calcium/calmodulin regulated protein phosphatase calcineurin [
66]. CARHSP1 is a paralogue of another cold shock protein PIPPin whose expression is limited to brain cells [
67,
68]. PIPPin is known to bind specifically to the 3′-UTR ends of both histone H1 and H3.3 mRNAs, encompassing the polyadenylation signal [
68]. Its role is implicated in the negative regulation of histone variant synthesis in the developing brain [
68]. PIPPin also interacts with other RNA binding proteins such as hnRNP A1, hnRNP K, and YB-1 [
69].
A further member of human cold shock protein is known as Unr (upstream of N-ras). It was first described as upstream of N-ras and initially identified as a regulator of N-ras expression [
70‐
73]
. Later it was found that Unr encodes a protein that possesses 5 CSDs, and is mainly expressed in the cytoplasm [
74,
75]. The gene was then renamed as CSD containing E1 (CSDE1). CSDE1 plays a key role in translational reprogramming by determining the fate of mRNAs by changing their stability and abundance [
76]. CSDE1 promotes and represses the translation of RNAs and also increases and decreases their abundance. Hence the role of CSDE1 is considered bidirectional [
76].
The final members of cold shock protein family in humans are denoted LIN28A or LIN28B, two highly related RNA binding proteins and proto-oncogenes [
77]. The role of LIN28 is to regulate translation of mRNAs that control developmental timing, pluripotency and metabolism [
78]. Besides, LIN28 is responsible for the repression of the let-7 microRNA biogenesis which is required for normal development and maintainence of the pluripotent state of cells [
79‐
81].
Cold shock proteins in plants
Cold shock proteins play pleiotropic functions in plants ranging from acquiring freezing tolerance to regulating embryo development, flowering time and fruit development [
82]. WCSP1 is the wheat cold shock protein that was the first functionally characterized plant cold shock protein [
83]. It possesses biochemical functions similar to bacterial cold shock proteins and is involved in cold adaptation. WCSP1 shows binding with both DNA and RNA and unwinds double-stranded nucleic acids in vitro and in vivo [
83‐
85]. In response to cold stress, there is upregulation of WCSP1 mRNA and increased expression of the corresponding protein in crown tissue during prolonged cold acclimation [
83]. Radkova et al
. serologically characterized the temporal and spatial distribution of the wheat CSD proteins with regard to plant development and cold adaptation [
86]. They identified 4 wheat cold shock protein genes through database analysis and classified into three classes based on their molecular masses and protein domain structures. Class I (20 kDa) and class II (23 kDa) wheat cold shock proteins were observed to accumulate in root and shoot meristematic tissues during vegetative growth. Protein expression of class I and class II wheat cold shock proteins remained high during flower and seed development. On the contrary, class III wheat cold shock protein (27 kDa) was detected only during seed development. In response to cold stress, wheat cold shock proteins accumulate in crown tissue which suggests their role in cold acclimation [
86].
Arabidopsis thaliana has 4 cold shock proteins 1–4 (AtCSP1-CSP4), that possess an N-terminal CSD. They all show binding with RNA, single and double-stranded DNA, and are able to unwind nucleic acid duplex. AtCSP3 (At2 g17870) is the only cold shock protein that is reported to be essential for the acquisition of freezing tolerance in
Arabidopsis [
88]. Overexpression of AtCSP3 confers freezing tolerance by regulating expression of stress-related genes whose roles in freezing tolerance are not known [
87]. Overexpression of AtCSP1 (CSDP1; At4g36020) is reported to delay seed germination under dehydration or salt stress conditions, whereas
AtCSP2 overexpression accelerates seed germination under salt stress [
88]. Juntawong et al. reported that AtCSP1 associates with polyribo-somes via an RNA-mediated interaction. AtCSP1 is implicated in selectively chaperoning mRNAs and improved translation of ribosomal protein mRNAs during cold stress [
89].
Plant cold shock proteins also regulate developmental processes. AtCSP2 is expressed many folds in meristematic tissues and ovules [
90‐
92], and regulates flowering transition, and flower and seed development [
90]. AtCSP4 (AtGRP2b; At2g21060) also plays an important role in development as
AtCSP4 overexpression leads to reduced silique length and induces embryo lethality [
93].
Chaikam and Karlson characterized the cold shock proteins in rice under different stress treatments and during various stages of development [
94]. The authors reported that two CSD proteins (OsCSP1 (Os02g0121100) and OsCSP2 (Os08g0129200)) in rice have nucleic acid binding activity and can complement a cold sensitive
E. coli strain [
94]. Expression of OsCSPs was found at a constant level during cold treatment that last over a period of several days. On the contrary, both OsCSP proteins and transcripts highly accumulated in reproductive tissues and tissues which exhibit meristematic activity [
94]. Thus, the role of OsCSPs may be more linked with developmental processes rather than with cold tolerance.
OsCSPs are maintained at a constant level subsequent to a cold treatment lasting over a period of several days.
A time-coursed study through various stages of rice development confirmed that both OsCSP proteins and transcripts are highly accumulated in reproductive tissues and tissues which exhibit meristematic activity. CSP1 associates with polyribosomes (polysomes) via an RNA-mediated interaction.
Conclusions
This review highlights the significance of cold shock proteins in several organisms and sheds new light on the unexplored cold shock protein member of P. falciparum. Functional diversity of cold shock protein family is entailed by discussing their known roles in gene expression regulation, cold adaptation, disease progression, and developmental processes such as flowering transition and flower and seed development. In silico work described here provide structural information of PfCoSP and hints its functional role particularly in regulating gene expression at gametocyte stages. However, the presence and structure–function characterization of PfCoSP needs to be confirmed in vivo and in vitro to conclude its role in gametocytogenesis and also at other stages of parasite life cycle. Future studies should aim at the structural and functional characterization of PfCoSP to understand its pivotal role in malaria parasites. Since cold shock proteins are verifiable targets for therapeutic intervention, the work described here, along with future studies on PfCoSP, may help with strategies aimed at targeting this protein directly for the development of anti-malarials and transmission-blocking vaccines.
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