Monthly Archives: February 2018

Gunter Blobel: Nobel Laureate Best known For the Signal Hypothesis of Protein Targeting

Günter Blobel, recipient of the 1999 Nobel Prize in Physiology or Medicine, died on February 18, 2018. Blobel is best known for his groundbreaking studies on an issue of fundamental importance to cell biology and virology—how newly synthesized proteins reach their correct location within or outside the cell.

Gunter Blobel (4)

In Blobel’s words: “Concurrently with or shortly after their synthesis on ribosomes, numerous specific proteins are unidirectionally translocated across or asymmetrically integrated into distinct cellular membranes. (1).” How this might be realized, now known as the “signal hypothesis,” was first put forward in 1971 by Blobel and colleague David Sabatini.  The central feature of the hypothesis is “the occurrence of a unique sequence of codons, located immediately to the right of the initiation codon, which is present only in those mRNAs whose translation products are to be transported across (my note: or inserted into) a membrane (2).”

Blobel and postdoc Bernhard Dobberstein (one of the impressive cell biologists trained by Blobel) confirmed the hypothesis in a classic experiment based on an earlier observation by Cesar Milstein. Specifically, Milstein’s group found that proteins, produced by translation of immunoglobin light chain mRNA, in a cell-free translation system, contained about 20 amino acids at their N-terminal end that are not present on immunoglobin light chains that are secreted from cells. Blobel and Dobberstein confirmed that cell-free translation (on free ribosomes) of immunoglobin light chain mRNA indeed yields the larger form of the protein. However, when membranes were added to the system, the light chains that were synthesized were the same size as normally secreted light chains. What’s more, the ribosomes synthesizing the light chains were bound to the membranes, and the new light chains were resistant to digestion by added proteases, indicating that they had been secreted into microsomes (2).

The overall scheme is that those nascent proteins, which are destined for secretion, or insertion into membranes, contain a short signal sequence at their N-terminus, which causes their ribosome to attach to a membrane. The signal sequence is removed by a signal peptidase as the growing polypeptide passes through a channel in the membrane. Proteins destined to be transmembrane proteins also contain a hydrophobic stop-transfer sequence, which anchors them in the membrane. Note that important details of this fundamental cellular pathway were revealed by Blobel’s analysis of the strategy by which sindbis virus generates its envelope glycoproteins (3). [For a detailed review of this key step in the replication cycles of enveloped viruses, see Chapters 7 and 8 of L. Norkin, Virology: Molecular Biology and Pathogenesis, ASM Press, 2010.]

Gunter Blobel was born in May 1936, in the Silesian village of Waltersdorf, then in eastern Germany, and later a part of Poland. He is another of the scientists featured on the blog whose life was impacted by events of the Second World War. Others include Max Delbruck, Francois Jacob, Jacques Monod, Andre Lwoff, the Wollmans (Eugene, Elizabeth, and Elie), Renato Dulbecco, Harald zur Hausen, and George Klein.

In Blobel’s words (4): “1945 was also a turning point in my life. Until then my childhood was a perfect 19th century idyll. In the cold and snow-rich Silesian winters there were hour-long rides on Sundays in horse-drawn sleighs to my maternal grandparent’s farm to have lunch and to spend the afternoon. The house was a magnificent 18th century manor house in the nearby Altgabel with a great hall that was decorated with hunting trophies. In the summer, of course, horse-drawn landauers were used as means of transportation. The way to school was a long one. We went there on foot and as a pack, usually consisting of one or two of my seven brothers and sisters and of children from neighboring houses.

At the end of January 1945, we had to flee from the advancing Russian Red Army. My father, a veterinarian stayed behind for a few more days and left only hours before the Red Army moved in. My fourteen-year-old brother, Reiner, drove my mother, my youngest brother, an older brother, the two younger sisters and me in a small automobile to relatives west of Dresden in Saxony. On the way there we drove through Dresden. We entered the city from the eastern hills. Its many spires and the magnificent cupola of the Frauenkirche (die Steinerne Glocke, the Stone Bell) were a magnificent sight even for the untrained eye of a child. Driving through Dresden, I still remember the many palaces, happily decorated with cherubs and other symbols of the baroque era. The city made an indelible impression on me. Only a few days, later, on February 13, 1945, we saw from a distance of about 30 kilometers a fire-lit, red night sky reflecting the raging firestorm that destroyed this great jewel of a city in one of the most catastrophic bombing attacks of World War II. It was a very sad and unforgettable day for me.

The months before and after the end of World War II were chaotic and miserable. None of my relatives had enough space to accommodate our large family leaving us divided among several relatives in different villages. There was no communication and little food. On September 9, 1945, we learned of the death of my beautiful oldest sister Ruth who, at age 19, was killed in an air raid on a train she was travelling in on April 10, 1945. She was buried in a mass grave near the site of the attack in Schwandorf, Bavaria. Ruth was born when my mother was just 20. The two had a sisterly relationship. My mother grieved over Ruth’s death until the end of her own life.

Fortunately, things took a turn for the better, when my father was able to continue his veterinarian practice in the charming medieval Saxon town of Freiberg. Most members of our family were reunited there by 1947. We lived in a nice villa surrounded by a large garden on the edge of town. My way to school was along the old medieval city wall. For only 40,000 inhabitants, Freiberg had a rich cultural life with a 175-year-old theater. Most impressive were the musical performances in the magnificent gothic cathedral, the Dom, with the splendid great Silbermann organ. Each week Bach cantatas were performed. The great choral works of Bach, Mozart and Haydn were regularly performed and at the highest artistic level at the major religious holidays. I even participated in singing in the cantus firmus of Bach’s Matthäus Passion. So, it was almost like a 19th century idyll again, this time in a small medieval town instead of a country village.

However, there was now the ever more oppressive regime of East Germany to deal with on a daily basis. When I graduated from high school in 1954 I was not allowed to continue my education at a university because I was considered a member of the “capitalist” classes. [My note:, Gunter was labeled “a member of the capitalist classes,” and barred from universities, for refusing to join a Communist youth group.] Fortunately, at that time, i.e., before the Berlin Wall, it was possible to escape and to travel freely to West Germany. So, on August 28, Goethe’s birthday, I left Freiberg for Frankfurt on the Main in West Germany. The train left in the morning and in the afternoon, it passed Weimar, where Goethe spent most of his life, and then Eisenach, where Bach was born and in the evening it arrived in Frankfurt, Goethe’s birthplace.”

Several paragraphs later:

“In 1994, I founded Friends of Dresden, Inc., a charitable organization, with the goal to raise funds in the U.S. to help rebuild the Frauenkirche in Dresden. The rebuilding of many of the historic monuments of Dresden is one of the most exciting consequences of German reunification and the liberation from communism. It is a childhood dream come true.

It was one of the great pleasures of my life to donate the entire sum of the Nobel Prize [my note: $960,000], in memory of my sister Ruth Blobel, to the restoration of Dresden, to the rebuilding of the Frauenkirche and the building of a new synagogue. This donation also serves to express my gratitude to my fellow Saxons. They received us with open arms when we had to flee Silesia. I spent a wonderful period of my life there and they gave me a thorough and valuable education. A few thousand dollars will also be donated for the restoration of an old baroque church in Fubine/Piemonte/ltaly, the home town of my wife’s father, Sebastanio Maioglio. We have spent many happy summers there in the parental home of my wife.” [My notes: the Frauenkirche was destroyed by the Allied bombing, and the Nazis destroyed the synagogue was in 1938. Globel also took up the rebuilding of the Paulinerkirche, the university church of the University of Leipzig, which had been blown up by the communist regime of East Germany in 1968.]

Blobel earned a degree in medicine from the University of Tubingen in 1960. However, after his internship in Germany, he decided that as a doctor he was merely treating symptoms, and that to get at causes he must turn to research. Blobel earned a doctorate in oncology from the University of Wisconsin in 1967. Next, he was a postdoctoral fellow at the Rockefeller Institute, under Nobel laureate, George Palade, and eventually became a full professor at the Institute in 1976, where he remained active until his recent death.

References:

  1. Blobel, G. (1980). Intracellular protein topogenesis, Proc Natl Acad Sci USA, 77:1496-1500.
  2. Blobel, G., and B. Dobberstein. (1975). Transfer of proteins across membranes. 1. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma. J Cell Biol, 67:835-851.
  3. Bonatti, S., R. Cancedda, and G. Blobel (1979). Membrane biogenesis. In vitro cleavage, core glycosylation, and integration into microsomal membranes of sindbis virus glycoproteins. J Cell Biol, 80:219-230. DOI: 10.1083/jcb.80.1.219
  4. “Günter Blobel – Biographical”. Nobelprize.org. Nobel Media AB 2014. Web. 21 Feb 2018. <http://www.nobelprize.org/nobel_prizes/medicine/laureates/1999/blobel-bio.html&gt;
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Knowledge Gaps in the Pursuit of a Universal Influenza Vaccine

Seasonal influenza outbreaks cause between 250,000 to 500,000 deaths word-wide each year (according to 2008 WHO estimates).  What’s more, unpredictable pandemics, of which there were four in the 20th and the 21st centuries (1918, 1957, 1968, and 2009) pose a still greater threat. The worst of these pandemics, the 1918 Spanish flu outbreak, claimed an estimated 50 to 100 million lives globally (according to 2014 WHO estimates).

The human population was “only” 1.9 billion individuals during the 1918 pandemic, whereas there now are about 8 billion people inhabiting our planet. Thus, a future pandemic might be far more catastrophic than the 1918 episode. Moreover, since pandemic strains are derived in part from zoonotic influenza viruses, the constant rise in livestock numbers, intensive farming, and the numbers of animals being transported around the world, combine to facilitate the genetic mixing and evolution of influenza viruses, and the chance of an animal influenza virus becoming able to jump to humans and causing a pandemic.

Unlike vaccines against other viruses (e.g., measles), the seasonal flu vaccine needs to be updated each year to keep up with the antigenic changes that continually occur in influenza viruses. This creates several problems, the first of which is that individuals need to be re-vaccinated each year. And, since it takes months to produce a vaccine, when the updated vaccine is at last ready, it may not be a particularly good match against the new season’s strains. The current vaccine is only about 30 percent effective, which accounts at least in part for the unusually severe flu season we are currently experiencing. And while the efficacy of the current vaccine may be atypically low, even in good years the match is less than optimal.

Pandemics present a much greater challenge to vaccine makers, since pandemics may be vastly more severe than seasonal outbreaks, and since an entirely new vaccine is needed against pandemic viruses. That latter is so because, as noted, pandemic strains are derived in part from zoonotic influenza viruses, which by-and-large are antigenically distinct from strains already circulating in the human population, that humans already express immunity against. Consider the example of the 2009 pandemic. Because an entirely new vaccine was needed to meet the threat of the pandemic virus, the vaccine was not available until after the first wave of infection had already occurred. Fortunately, the 2009 pandemic virus was relatively mild.

A “universal” flu vaccine, that could provide lifelong protection against all seasonal strains of influenza, as well as provide protection against a pandemic virus, would be a most crucial and significant breakthrough. A major international workshop, entitled “Pathway to a Universal Influenza Vaccine,” was convened June 28 and 29, 2017, by the U.S. National Institute of Allergy and Infectious Diseases, to identify gaps in our knowledge that need to be addressed to develop such a vaccine (1).

Participants noted shortcomings in our understanding of the epidemiology, transmission, natural history, and pathogenesis of influenza. Among the issues specifically mentioned: “influenza surveillance is lacking in certain regions of the developing world and globally in certain high-risk groups… gaps in knowledge include the relationship between symptoms, viral shedding and transmission, as well as the level of protection needed to interrupt transmission.”

The host factors that influence influenza disease severity were also acknowledged to be poorly understood. To that point, participants addressed the need to better understand how pre-existing immunity—which might result from multiple natural influenza infections, as well as from repeated vaccinations—might affect how that person’s immune response will respond to future infections and, importantly, how past exposures might affect the efficacy of a vaccine. As stated in the meeting report: “Recent data provide strong epidemiologic evidence that infection with the influenza strain circulating during one’s childhood elicits a lifelong immunologic imprint that impacts responses to novel strains and can help protect against unfamiliar HA subtypes from the same phylogenetic group as the original infecting virus…The potential consequences of imprinting infants with vaccines versus natural exposure need to be carefully assessed.” [The HA protein is the so-called hemagglutinin, which serves the virus as its attachment protein.]

Participants also noted gaps in our understanding of the underlying B and T cell immune mechanisms that are induced by both natural infections and vaccinations. Further study of these responses was recommended so that we might be better able to stimulate them.

As might be expected, it is singularly important to identify the antigens that might be the most promising targets of a universal vaccine. To that point: “…most areas of the HA head are subject to antigenic change, and therefore unlikely to yield a broadly protective immune response. Neutralizing antibody responses to conserved regions such as the HA stalk, and non-neutralizing antibodies such as those directed at the neuraminidase (NA), and matrix 2 ectodomain, merit further study…The importance of each site may differ for pandemic versus seasonal influenza.” [The HA head region binds to receptors on the cell surface. After the bound virus is taken into the cell by receptor-mediated endocytosis, the low pH within endosomes triggers a conformational rearrangement of the HA stalk region, which mediates fusion of the viral envelope with the endosomal membrane, thereby releasing the viral cores into the cytosol. Since mutations within the stalk region might disrupt its membrane fusion function, such mutations are generally selected against. Researchers have recently had success developing antibodies that target the neuraminidase protein at the viral surface.]

Participants acknowledged that animal models play an important role in influenza research, especially when studying pandemic viruses. Nonetheless, “Animal models have limitations including the inability to mimic the human experience regarding genetic background, lifetime exposure to natural influenza infection or vaccine, viral susceptibility…and determinants of immune response and protection.” Thus, the participants noted that “a human a challenge model will be a crucial tool for vaccine development, as it can help answer fundamental questions about influenza immunity and serve as a mechanism for rapidly testing the efficacy of new products…Expansion of this resource should be a top priority…”

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Day two of the workshop consisted of a rapporteur session on key conclusions, chaired by David Baltimore and Anthony S. Fauci (1)

Participants agreed that a robust collaboration between government agencies, academia, and industry would be needed to translate the fruits of basic research into a universal influenza vaccine. To that point, a January 24, 2018 editorial in Nature (doi: 10.1038/d41586-018-01070-w) asserted: “…advocates rightly argue that the research and development of a universal flu vaccine — ultimately the only effective defense against future pandemics — merits a program equivalent in scale to the Manhattan Project.” Yet the US government last year invested just $75 million on universal flu vaccine research and development.

    1. Paules C.I., D. Marston, R. W. Eisinger, D. Baltimore, and A. S.Fauci. (2017), Meeting Report: The Pathway to a Universal Influenza Vaccine. Immunity 47: 599-603. https://doi.org/10.1016/j.immuni.2017.09.007