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Today, a research result [1] reported in the journal "Science" has once again aroused the attention of the academia and the public to the new coronavirus.
The research team led by Professor Rudolf Jaenisch, a famous biology and geneticist at MIT Whitehead Institute, published an unexpected research result on the preprint platform bioRxiv [2].
After analyzing the historical research data of the new coronavirus, Jaenisch discovered that there is a phenomenon of fusion of the new coronavirus fragment with human genomic DNA.
They subsequently confirmed through in vitro cell line studies that part of the nucleic acid sequence of the new coronavirus can be integrated into the human genome based on the widespread LINE-1 retrotransposon in the human body or the HIV-1 reverse transcriptase in HIV-infected persons.
in.
More importantly, the new coronavirus fragments integrated into the human genome can be transcribed normally.
At present, the impact of this feature of the new coronavirus on the human body is still unknown.
However, this finding explains to a certain extent the phenomenon of rejuvenation of patients after rehabilitation.
▲ Screenshots of this research Earlier this year, we often saw reports that nucleic acid tests of survivors of new coronary pneumonia suddenly or continued to be positive [3-6].
Although there have been several reports of re-infection after recovering from new coronavirus infection some time ago [7], a cohort study of strictly quarantined recovered patients has shown that Fuyang cases are not caused by re-infection [8, 9].
What is even more strange is that no virus with replication ability has been isolated in the above-mentioned PCR-positive recovered patients [3-5].
This strange phenomenon of rejuvenation has always lacked a reasonable explanation.
Jaenisch and his colleagues speculate that as a positive-strand RNA virus, the RNA of the new coronavirus may be reverse-transcribed into DNA and then integrated into the human genome.
The transcriptionally integrated DNA copy may be the cause of the positive PCR test.
.
In fact, scientists have observed the activity of endogenous reverse transcriptase in human cells, and have proven that the products of reverse transcription can be integrated into the human genome [10, 11].
For example, the LINE-1 retrotransposon, which accounts for about 17% of the human genome DNA, is a ready-made autonomous retrotransposon [10, 12].
In addition, for HIV-infected people, the HIV-1 reverse transcriptase in their bodies may also be an accomplice of the new coronavirus.
▲ Professor Rudolf Jaenisch Since the new coronavirus has a theoretical basis for integrating genetic material into the human genome, it needs to find evidence support from the real world.
The Jaenisch team first analyzed the published RNA sequencing data of cells infected with the new coronavirus to find evidence of the presence of integrated transcripts.
Based on the data obtained from nearly 10 studies [13-15], they really found a large number of integrated sequences composed of host genome and viral nucleic acid sequences in a variety of cell types.
▲Analysis of historical research data Generally speaking, integrated sequences generally account for 0.
004%-0.
14% of the total number of new coronavirus sequences in each sample.
Among them, bronchoalveolar lavage fluid cells from patients with severe new coronary pneumonia account for the largest proportion, 69.
24%.
Moreover, most of the new coronavirus part of the integrated sequence comes from the nucleocapsid (N), which is consistent with the nucleocapsid RNA being the most abundant RNA in the new coronavirus subgenome [16], so it is most likely to be reverse transcription And integration goals.
Jaenisch and his colleagues believe that the above data supports the hypothesis that the new coronavirus can integrate into the genome of infected cells and produce integrated virus-cell transcripts.
▲ New coronavirus-NIAID As we have already introduced in the previous article, the new coronavirus may be integrated into the human genome with the help of LINE-1 retrotransposon or HIV-1 reverse transcriptase.
To confirm this conjecture, Jaenisch and his colleagues overexpressed human LINE-1 or HIV-1 reverse transcriptase in HEK293T cells, and then infected the transduced cells with the new coronavirus.
Two days after infection, the virus sequence of the cells was detected by PCR or fluorescence in situ hybridization (FISH).
Subsequently, the researchers obtained 4 PCR primer combinations targeting the nucleocapsid from the WHO, and detected the presence of the new coronavirus nucleocapsid sequence in the genomic DNA of the cell, and can clone the full-length nucleocapsid DNA.
Sanger sequencing results This is also confirmed.
The detection result of the fluorescently labeled probe showed that the nucleocapsid sequence signal did exist in the nucleus.
Moreover, in cells infected with the new coronavirus but not overexpressing LINE-1 retrotransposon or HIV-1 reverse transcriptase, nucleocapsid signals were also detected in the nuclear region, although the frequency was low (about 10%).
), which also means that the integration of the new coronavirus nucleocapsid RNA is achieved through the endogenous reverse transcriptase activity of the cell.
▲Is there any effect of LINE-1 on integration However, there is still a real problem.
Although LINE-1 retrotransposons account for a relatively high proportion of genomic DNA, their activity is very low, about 5000 One is active [10,12].
Since the activity of the LINE-1 retrotransposon is so low, is the integration of the new coronavirus into the human genome really related to it? In order to solve the above problems, Jaenisch and his colleagues also called up the published RNA sequencing data of cells infected with the new coronavirus.
In these data, they found that the expression of LINE-1 was significantly up-regulated and correlated with the abundance of integrated sequences.
Cell studies have shown that when infected with the new coronavirus, the expression of LINE-1 in CALU3 cells is up-regulated by about 3-4 times.
As for the reason why the new coronavirus infection promotes the up-regulation of LINE-1 expression, it is the familiar cytokine.
Jaenisch and his colleagues cultured bone marrow cells, microglia or CAR-T cells in a conditioned medium containing cytokines.
PCR analysis found that the expression of endogenous LINE-1 was up-regulated by about 2-3 times. The new coronavirus infection will induce the production of cytokines, and even induce a cytokine storm in severe cases [17].
▲ New coronavirus-NIAID In general, based on all the above data, the Jaenisch team believes that their research results show that the cytokine pressure induced by the new coronavirus infection induces the expression of LINE-1, which in turn promotes the new coronavirus to fragment genetic material.
Integrate into the human cell genome.
However, the new coronavirus sequence inserted into the human genome by reverse transcription is likely to be a subgenomic fragment, because the integrated sequence is mostly enriched in the nucleocapsid sequence, which also excludes the possibility of infectious virus production.
Simply put, the new coronavirus fragment integrated into the human genome should not produce a complete virus with infectious ability.
However, because the new coronavirus fragments integrated into the human genome can be transcribed, this provides an explanation for the phenomenon of rejuvenation in many recovered patients.
A question that Jaenisch and his colleagues are very concerned about is whether these new coronavirus sequence fragments integrated into the human genome can express viral antigens.
If possible, how does this affect the course and treatment of the disease?
Of course, the above influence also depends on the position of the new coronavirus fragment inserted in the genome, which has been confirmed in HIV-infected persons [18].
Therefore, in-depth study of the integration sites of the new coronavirus in the patient's genomic DNA and the correlation between these sites and the severity of the disease may help clarify some clinical problems.
▲ The new crown virus-NIAID has received mixed reviews from the academic community for the research results of the Jaenisch team.
Some people even made severe criticisms, believing that this study is incomplete and did not determine where the new coronavirus is inserted into the human genome.
It cannot be ruled out that it is the problem of PCR amplification itself.
Therefore, it is impossible to conclude that the new coronavirus fragment will be inserted into the human genome [2 ]. Some people were shocked by the Jaenisch team’s publication of this article at this point in time, because now coincides with the launch of mRNA vaccines, this research will cause public concern about mRNA vaccines, and they even worry that this article will be used by anti-vaccine people[ 2].
In addition to critical voices, there are also some positive voices.
For example, Robert Gallo, a well-known virologist and head of the Institute of Human Virology at the University of Maryland School of Medicine, said in an interview with "Science" that although this is not a complete story, he still likes the study and guessed that it should be correct.
1].
David Baltimore, a virologist at the California Institute of Technology who won the Nobel Prize in Physiology or Medicine for the discovery of reverse transcriptase, described the new work as "impressive" and the results of the study were "unexpected.
" Baltimore also believes that "this work Many interesting questions were raised".
Finally, in any case, as Baltimore said, this work raises many questions.
The editor talks about the development of the GIST field in the past two decades, which is a classic epitome of the development of molecular targeted therapy for cancer.
In the past three months, the Singularity team has also tried to overlook the panorama of the development of the GIST field from a higher perspective, and carefully created a GIST panorama course for everyone: "8 Lectures on GIST Academic Frontiers".
The 8-lecture course covers the latest developments in GIST treatment, deeply analyzes the mechanism of action of different targeted drugs, and systematically sorts out important global research in the GIST field, allowing you to gain insights into the cutting-edge academic progress of GIST within 100 minutes.
The research team led by Professor Rudolf Jaenisch, a famous biology and geneticist at MIT Whitehead Institute, published an unexpected research result on the preprint platform bioRxiv [2].
After analyzing the historical research data of the new coronavirus, Jaenisch discovered that there is a phenomenon of fusion of the new coronavirus fragment with human genomic DNA.
They subsequently confirmed through in vitro cell line studies that part of the nucleic acid sequence of the new coronavirus can be integrated into the human genome based on the widespread LINE-1 retrotransposon in the human body or the HIV-1 reverse transcriptase in HIV-infected persons.
in.
More importantly, the new coronavirus fragments integrated into the human genome can be transcribed normally.
At present, the impact of this feature of the new coronavirus on the human body is still unknown.
However, this finding explains to a certain extent the phenomenon of rejuvenation of patients after rehabilitation.
▲ Screenshots of this research Earlier this year, we often saw reports that nucleic acid tests of survivors of new coronary pneumonia suddenly or continued to be positive [3-6].
Although there have been several reports of re-infection after recovering from new coronavirus infection some time ago [7], a cohort study of strictly quarantined recovered patients has shown that Fuyang cases are not caused by re-infection [8, 9].
What is even more strange is that no virus with replication ability has been isolated in the above-mentioned PCR-positive recovered patients [3-5].
This strange phenomenon of rejuvenation has always lacked a reasonable explanation.
Jaenisch and his colleagues speculate that as a positive-strand RNA virus, the RNA of the new coronavirus may be reverse-transcribed into DNA and then integrated into the human genome.
The transcriptionally integrated DNA copy may be the cause of the positive PCR test.
.
In fact, scientists have observed the activity of endogenous reverse transcriptase in human cells, and have proven that the products of reverse transcription can be integrated into the human genome [10, 11].
For example, the LINE-1 retrotransposon, which accounts for about 17% of the human genome DNA, is a ready-made autonomous retrotransposon [10, 12].
In addition, for HIV-infected people, the HIV-1 reverse transcriptase in their bodies may also be an accomplice of the new coronavirus.
▲ Professor Rudolf Jaenisch Since the new coronavirus has a theoretical basis for integrating genetic material into the human genome, it needs to find evidence support from the real world.
The Jaenisch team first analyzed the published RNA sequencing data of cells infected with the new coronavirus to find evidence of the presence of integrated transcripts.
Based on the data obtained from nearly 10 studies [13-15], they really found a large number of integrated sequences composed of host genome and viral nucleic acid sequences in a variety of cell types.
▲Analysis of historical research data Generally speaking, integrated sequences generally account for 0.
004%-0.
14% of the total number of new coronavirus sequences in each sample.
Among them, bronchoalveolar lavage fluid cells from patients with severe new coronary pneumonia account for the largest proportion, 69.
24%.
Moreover, most of the new coronavirus part of the integrated sequence comes from the nucleocapsid (N), which is consistent with the nucleocapsid RNA being the most abundant RNA in the new coronavirus subgenome [16], so it is most likely to be reverse transcription And integration goals.
Jaenisch and his colleagues believe that the above data supports the hypothesis that the new coronavirus can integrate into the genome of infected cells and produce integrated virus-cell transcripts.
▲ New coronavirus-NIAID As we have already introduced in the previous article, the new coronavirus may be integrated into the human genome with the help of LINE-1 retrotransposon or HIV-1 reverse transcriptase.
To confirm this conjecture, Jaenisch and his colleagues overexpressed human LINE-1 or HIV-1 reverse transcriptase in HEK293T cells, and then infected the transduced cells with the new coronavirus.
Two days after infection, the virus sequence of the cells was detected by PCR or fluorescence in situ hybridization (FISH).
Subsequently, the researchers obtained 4 PCR primer combinations targeting the nucleocapsid from the WHO, and detected the presence of the new coronavirus nucleocapsid sequence in the genomic DNA of the cell, and can clone the full-length nucleocapsid DNA.
Sanger sequencing results This is also confirmed.
The detection result of the fluorescently labeled probe showed that the nucleocapsid sequence signal did exist in the nucleus.
Moreover, in cells infected with the new coronavirus but not overexpressing LINE-1 retrotransposon or HIV-1 reverse transcriptase, nucleocapsid signals were also detected in the nuclear region, although the frequency was low (about 10%).
), which also means that the integration of the new coronavirus nucleocapsid RNA is achieved through the endogenous reverse transcriptase activity of the cell.
▲Is there any effect of LINE-1 on integration However, there is still a real problem.
Although LINE-1 retrotransposons account for a relatively high proportion of genomic DNA, their activity is very low, about 5000 One is active [10,12].
Since the activity of the LINE-1 retrotransposon is so low, is the integration of the new coronavirus into the human genome really related to it? In order to solve the above problems, Jaenisch and his colleagues also called up the published RNA sequencing data of cells infected with the new coronavirus.
In these data, they found that the expression of LINE-1 was significantly up-regulated and correlated with the abundance of integrated sequences.
Cell studies have shown that when infected with the new coronavirus, the expression of LINE-1 in CALU3 cells is up-regulated by about 3-4 times.
As for the reason why the new coronavirus infection promotes the up-regulation of LINE-1 expression, it is the familiar cytokine.
Jaenisch and his colleagues cultured bone marrow cells, microglia or CAR-T cells in a conditioned medium containing cytokines.
PCR analysis found that the expression of endogenous LINE-1 was up-regulated by about 2-3 times. The new coronavirus infection will induce the production of cytokines, and even induce a cytokine storm in severe cases [17].
▲ New coronavirus-NIAID In general, based on all the above data, the Jaenisch team believes that their research results show that the cytokine pressure induced by the new coronavirus infection induces the expression of LINE-1, which in turn promotes the new coronavirus to fragment genetic material.
Integrate into the human cell genome.
However, the new coronavirus sequence inserted into the human genome by reverse transcription is likely to be a subgenomic fragment, because the integrated sequence is mostly enriched in the nucleocapsid sequence, which also excludes the possibility of infectious virus production.
Simply put, the new coronavirus fragment integrated into the human genome should not produce a complete virus with infectious ability.
However, because the new coronavirus fragments integrated into the human genome can be transcribed, this provides an explanation for the phenomenon of rejuvenation in many recovered patients.
A question that Jaenisch and his colleagues are very concerned about is whether these new coronavirus sequence fragments integrated into the human genome can express viral antigens.
If possible, how does this affect the course and treatment of the disease?
Of course, the above influence also depends on the position of the new coronavirus fragment inserted in the genome, which has been confirmed in HIV-infected persons [18].
Therefore, in-depth study of the integration sites of the new coronavirus in the patient's genomic DNA and the correlation between these sites and the severity of the disease may help clarify some clinical problems.
▲ The new crown virus-NIAID has received mixed reviews from the academic community for the research results of the Jaenisch team.
Some people even made severe criticisms, believing that this study is incomplete and did not determine where the new coronavirus is inserted into the human genome.
It cannot be ruled out that it is the problem of PCR amplification itself.
Therefore, it is impossible to conclude that the new coronavirus fragment will be inserted into the human genome [2 ]. Some people were shocked by the Jaenisch team’s publication of this article at this point in time, because now coincides with the launch of mRNA vaccines, this research will cause public concern about mRNA vaccines, and they even worry that this article will be used by anti-vaccine people[ 2].
In addition to critical voices, there are also some positive voices.
For example, Robert Gallo, a well-known virologist and head of the Institute of Human Virology at the University of Maryland School of Medicine, said in an interview with "Science" that although this is not a complete story, he still likes the study and guessed that it should be correct.
1].
David Baltimore, a virologist at the California Institute of Technology who won the Nobel Prize in Physiology or Medicine for the discovery of reverse transcriptase, described the new work as "impressive" and the results of the study were "unexpected.
" Baltimore also believes that "this work Many interesting questions were raised".
Finally, in any case, as Baltimore said, this work raises many questions.
The editor talks about the development of the GIST field in the past two decades, which is a classic epitome of the development of molecular targeted therapy for cancer.
In the past three months, the Singularity team has also tried to overlook the panorama of the development of the GIST field from a higher perspective, and carefully created a GIST panorama course for everyone: "8 Lectures on GIST Academic Frontiers".
The 8-lecture course covers the latest developments in GIST treatment, deeply analyzes the mechanism of action of different targeted drugs, and systematically sorts out important global research in the GIST field, allowing you to gain insights into the cutting-edge academic progress of GIST within 100 minutes.
