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\DOI{10.5802/crbiol.141}
\datereceived{2023-10-24}
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\dateposted{2023-12-19}
\datepublished{2024-03-29}
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%\makeatletter
%\def\TITREspecial{\relax}
%\def\cdr@specialtitle@english{A tribute to Fran\c{c}ois Gros, a founding father of molecular biology}
%\def\cdr@specialtitle@french{Un hommage \`a Fran\c{c}ois Gros, un des p\`eres de la biologie mol\'eculaire}
%\makeatother

\title{Memories of Professor Fran\c{c}ois Gros}

\alttitle{Souvenirs du professeur Fran\c{c}ois Gros}

\author{\firstname{Shin'ichi} \lastname{Takeda}\CDRorcid{0000-0003-4780-6290}}
\address{National Institute of Neuroscience,
National Center of Neurology and Psychiatry,
Japan}
\email{takeda@ncnp.go.jp}

\keywords{\kwd{Fran\c{c}ois Gros}
\kwd{Memories}
\kwd{Muscle biology}
\kwd{Muscular dystrophy}}

\altkeywords{\kwd{Fran\c{c}ois Gros}
\kwd{Souvenirs}
\kwd{Biologie musculaire}
\kwd{Dystrophie musculaire}}

\begin{abstract} 
I joined the laboratory of Professor Francois Gros in 1987 and worked there
as a postdoc with Robert Whalen until 1992. I recount the research we
carried out and mention that of the other scientists also working on
skeletal muscle on the 6th floor of the Molecular Biology Department of
the Institut Pasteur at that time. I then present my subsequent
research when I returned to Japan. I pay tribute to the influence of
Professor Gros and to his support in establishing Japanese/French meetings
on muscle biology and muscular dystrophy. I also invoke personal
memories of Robert Whalen and Margaret Buckingham and remember the
occasions when I returned to Paris to honour Fran\c{c}ois Gros.
\end{abstract}

\begin{altabstract} 
J'ai rejoint le laboratoire du professeur Fran\c{c}ois Gros en 1987 et
j'y ai travaill\'{e} en tant que postdoc avec Robert Whalen jusqu'en
1992. Je raconte les recherches que nous avons men\'{e}es et je
mentionne celles des autres scientifiques qui travaillaient
\'{e}galement sur le muscle squelettique au $6^{\rm e}$
\'{e}tage du D\'{e}partement de biologie mol\'{e}culaire de l'Institut
Pasteur \`{a} cette \'{e}poque. Je pr\'{e}sente ensuite les recherches
ult\'{e}rieures que j'ai men\'{e}es de retour au Japon. Je rends
hommage \`{a} l'influence du professeur Gros et \`{a} son soutien lors
de la mise en place de r\'{e}unions franco-japonaises autour de la
biologie musculaire et de la dystrophie musculaire. J'\'{e}voque
\'{e}galement des souvenirs personnels de Robert Whalen et de Margaret
Buckingham et je me rappelle les moments o\`{u} je suis retourn\'{e}
\`{a} Paris pour rendre hommage \`{a} Fran\c{c}ois Gros.
\end{altabstract}

\maketitle

\vspace*{8pt}

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\xsection{}
From the beginning of August 1987 to the end of August 1992, I was with
Professor Fran\c{c}ois Gros at the Institut Pasteur (IP) in Paris as a
postdoctoral fellow. My study abroad was made possible through the
recommendations of Professor Nobuo Yanagisawa, of the Third Department of
Internal Medicine (Neurology) at Shinshu University, School of
Medicine, the late Professor Setsuro Ebashi, and the late Professor Yoshiaki
Nonomura of the Department of Pharmacology at the University of Tokyo,
School of Medicine, who taught me during my graduate 
school days.

\section{On the 6th floor of the Molecular Biology Building in
IP}\label{sec1} 
I wonder what muscle biology looked like at the Institut Pasteur when I
studied there. Professor Fran\c{c}ois Gros, a disciple of Nobel Prize
winner Dr. Jacques Monod, contributed to the discovery of mRNA in the
1960s, then in the 1970s he became interested in higher-order
biological functions and decided to\unskip\break 
focus his research on muscle development
and differentiation. He set up a laboratory, Unit\'{e} de
Biochimie, on the 6th floor of the Molecular Biology Building in IP.
Professor F. Gros served as the director general of the Pasteur
Institute in the early 1980s, and in the late 1980s he served as a
special scientific advisor to President Fran\c{c}ois Mitterrand. When I
was in IP, there were three principal researchers on the 6th floor of
the building. One was Dr. Margaret Buckingham, who was already
independent and had her own laboratory. While also a professor at the
Coll\`{e}ge de France with a laboratory there, Professor F. Gros worked
with two principal investigators, Dr. Robert G. Whalen and Dr. Marc
Fiszman. At that time, Margaret focused on the development and
differentiation of skeletal muscle and mainly dealt with myosin light
chain (MLC) genes, Robert (Bob) dealt with postnatal muscle development
and myosin heavy chain (MyHC) genes, and Marc focused on the
relationship between muscle differentiation and splicing, especially of
Tropomyosin genes. Therefore, there was considerable complementary
between the scientific interests of the groups sharing 
the 6th floor. 

The biggest advances in skeletal muscle biology that occurred during my
five-year stay were the discovery of muscle regulatory factors (MRFs),
centered on the MyoD gene, and the discovery of dystrophin, which is
the cause of Duchenne muscular dystrophy (DMD). The former subject was
pursued on the 6th floor of the Molecular Biology Building, which was
recognized as one of the centers of research on the expression and role
of these factors. MRFs consist of four transcription factors (MyoD,
Myogenin, Myf5, and Mrf4), and Myogenin was in part discovered through
collaborative research with Dr. Woodring Wright~\cite{1}, working on
the 6th floor. An \textit{in situ} hybridization study on the
expression of MRFs was published in \textit{Nature} by Dr. David
Sassoon, Margaret's postdoctoral fellow~\cite{2}, and Shahragim
Tajbakhsh, another postdoc in her lab, created a Myf5 null
mouse~\cite{3}. Heterozygous Myf5 mice, in which the \textit{lacZ}
reporter gene has been inserted, are frequently used in muscle
development research to this day. Members of the laboratory of Dr.
Thomas Braun, who created the Myf5 null mouse one step ahead of
Shahragim, Dr. Giulio Cossu, who succeeded in identifying
mesoangioblast/pericyte stem cells, and Dr. Stefano Schiaffino, who
created the MyHC monoclonal antibodies that are still used around the
world, often visited Margaret or stayed in her lab for longer periods
of time.

\section{Unit\'{e} de Biochimie directed by Dr. Fran\c{c}ois
Gros}\label{sec2}
Under these favorable circumstances, I was able to participate in Bob's
research group with the support of Professor Fran\c{c}ois Gros. Bob
achieved a number of groundbreaking results from the late 1970s to the
1980s. These included the discovery of an embryonic myosin heavy 
chain~\cite{4}, and the discovery of the expression of a skeletal
muscle-type myosin light chain in the fetal  heart~\cite{5}. Then, he
found sequential expression of embryonic, neonatal, and adult types of
myosin heavy chains during developmental stages of rat skeletal 
muscle~\cite{6}. Furthermore, innervation, thought to be a key
regulatory mechanism controlling the expression of these isozymes, was
investigated in his lab, by using denervation procedures. He observed
that denervation does not interfere with the expression of the adult
type of myosin heavy  chain~\cite{7}. I will never forget that
Professor Fran\c{c}ois Gros gave a detailed summary of these studies
upon the retirement of Dr. Gillian Butler-Browne, who was a
co-researcher on many of them. As a postdoctoral fellow under Dr.
Robert G. Whalen, I worked hard on cloning the mouse MyHC IIB gene
promoter and elucidating its transcriptional regulatory mechanism, and
published several  papers~\cite{8,9,10}. Subsequent research did not
sufficiently clarify the transcriptional regulatory mechanism of MyHC
IIB gene, but in recent years we have been able to shed some light on
it through research on mice living on the space station. It involves a
group of transcription factors called Maf. There are three types of Maf
expressed in skeletal muscle, and when we created triple knockout mice
in which all of them were knocked out, the expression of MyHC IIB
disappeared, which revealed the regulatory mechanism from the
transcriptional factor side for the first  time~\cite{11}. Meanwhile,
Bob then ventured into a completely new field of biology. This was an
important attempt to create a vaccine by injecting into skeletal
muscle, as a plasmid, a part of the genes that make up the
virus~\cite{12}. The 2023 Nobel Prize in Physiology and Medicine was
awarded to two researchers who developed an mRNA vaccine against the
new type of coronavirus, but, although there is a difference between
DNA and RNA, Bob's attempt largely preceded their work. Therefore, I
think it deserves more recognition. The insight of Professor Fran\c{c}ois
Gros in realising the potential of Bob as a young researcher is also to
be praised.

\section{What I have done after I went back to Japan}\label{sec3}
I returned to Japan in 1992 and worked at the National Institute of
Neuroscience, National Center of Neurology and Psychiatry, NCNP, in
Kodaira City, Tokyo. What kind of work have I been able to accomplish
at the National Institute of Neuroscience since then? It is broadly
divided into four fields. The first was research to develop treatments
for muscular dystrophy, which I began after returning to 
Japan~\cite{13,14}. I have also focused my efforts on researching the
pathology of muscular dystrophy, with the development of a mouse model
of muscular dystrophy using a gene knockout  approach~\cite{15,16}. As
a further development, we were able to establish a colony of muscular
dystrophy dogs, which are medium-sized and have a more severe form of
the disease, thus providing a good animal  model~\cite{17}. The third
category of my research was in muscle cell biology, which can be seen
as a direct consequence of the research on skeletal muscle
differentiation conducted at IP. In 2007, we published an expression
analysis study using microarrays on muscle satellite cells, which are
one of the tissue stem cells of skeletal  muscle~\cite{18}.
Furthermore, in 2010, we showed that PDGFR$\rmalpha$-positive
mesenchymal progenitors exist in the mesenchyme of skeletal muscle, and
that they not only positively control muscle regeneration but also are
the origin of adipogenic cells and  fibrosis~\cite{19}. Based on these
studies, we are currently conducting research to induce muscle lineage
stem cells using iPS cells and apply them to regenerative medicine, but
this research has a long way to go. Finally, I gradually began
researching signal transduction in skeletal muscle. We were able to
demonstrate that neuronal nitric oxide synthase (nNOS) and nitric oxyde
play an important role not only in muscle  atrophy~\cite{20}, but also
in muscle hypertrophy~\cite{21}. In particular, the latter research has
uncovered the calcium signal transduction mechanism centered on the
sarcoplasmic reticulum (SR), making it possible to reconsider the
pathology of muscular dystrophy from the perspective of the calcium
control  mechanism~\cite{22}.

\section{The influence of Professor Fran\c{c}ois Gros on me}\label{sec4}
After returning to Japan, I began therapeutic research for muscular
dystrophy, but in carrying out this translational research, I needed to
study not only basic and clinical medicine, but also biochemistry, cell
biology, molecular genetics, and molecular biology. An integrated
understanding was necessary. At that time, unlike Japan, where research
tends to be carried out vertically in each field, France, where
cross-disciplinary exchanges are encouraged in a free atmosphere, was
an unforgettable experience for me, especially in the IP environment. I
recall that there was a visit from Japan to Professor Fran\c{c}ois Gros'
laboratory in the context of fostering a Center of Excellence (a
nationally representative research facility like IP), which later
became one of the pillars of scientific research policy in Japan.
Therefore, I came up with the idea of holding a workshop to connect
Japan and France, and on March 20, 1995, I was able to hold an initial
research meeting. The key participants in the meeting at IP 
(Figure~\ref{fig1}) were Dr. Robert G. Whalen, who was the head of the
team at IP, and Dr. Gillian Butler-Browne, who was about to move from
IP to the Institut de Myologie (IdM). On the Japanese side, Dr. Hideo
Sugita, President of NCNP (who unfortunately passed away in November
2019) agreed with the project and organised support for the travel and
event expenses, together with Dr. Tadayuki Ishihara, who was the vice
director of Higashi Saitama National Hospital (and later served as
director of Hakone National Hospital). Needless to say, we had the
support of Professor Fran\c{c}ois Gros. The day the research meeting
was held, the sarin gas attack on the Tokyo subway occurred, making it
an unforgettable day. As a result of this initial meeting, we were
subsequently able to hold an international Japan-France workshop on
muscle biology and muscular dystrophy between the two countries.

\begin{figure}
\includegraphics{fig01}
{\vspace*{-2pt}}
\caption{\label{fig1}A circular of the scientific meeting at the Institut Pasteur
on 20/03/1995. This meeting was the precursor to a series of
12~consecutive Japan-France international workshops for muscular
dystrophy.}
{\vspace*{-2pt}}
\end{figure}

To date, we have been able to hold this workshop 12 times, and there
are several reasons for this. One was the ``joy of being exposed to
science'', which allowed us to bring together and present remarkable
scientific results. Japan has a long history of biochemistry and cell
biology, while France excelled in the fields of molecular biology and
molecular genetics, which must have been a source of reciprocal
inspiration. Next, it was also important that the workshop provided an
opportunity for mutual exchange among researchers. Furthermore, the
success of the first workshop held in Tokyo in 1996, followed by the
second held in Paris the following year 1997, was a major factor in the
decision to continue. On the French side, Dr. Michel Fardeau, Dr.
Fernando Tom\'{e}, Dr. Pascal Guicheney, Dr. Marc Fiszman, Dr. Thomas
Voit, Dr. Gillian Butler-Browne, and Dr. Gis\`{e}le Bonne from IdM, and
on the Japanese side, the late Dr. Sugita, Dr. Eijiro Ozawa, the late
Dr. Kiichi Arahata, Dr. Ikuya Nonaka, Professor Makiko Osawa, and Dr.
Ichizo Nishino were major participants in this effort. It was only
possible to continue the international workshop to this day due to the
great contributions of these researchers and clinicians.

\section{Personal memories}\label{sec5} 
I would like to express my gratitude to these two benefactors. The
first person is Dr. Robert G. Whalen, who was the head of the IP team.
When I was working on transcriptional regulation of the murine MyHC IIB
gene, he said to me, ``You have directly treated patients with muscular
dystrophy as a clinician, and at the University of Tokyo, you studied
the biochemistry of muscle proteins. If you could acquire the approach
of molecular biology of skeletal muscle at the genetic level, then you
would be a unique person in this field of research.'' He continued to
give me research opportunities. I've never met a person as talented as
him. When I left IP, I will never forget the tears in his eyes as he
bid me farewell.

The second person was Dr. Margaret Buckingham, who was on the same
floor but in the laboratory next door and later became a member of the
French Academy of Sciences. Margaret created an opportunity for me to
get a position in Japan. In 1990, an international symposium entitled
``Muscle Development, Maintenance, and Contraction; Recent
Developments'' was held in Japan, sponsored by the Uehara Memorial Life
Science Foundation. Margaret was invited and gave an excellent lecture;
she mentioned to many Japanese researchers that a researcher named
Shin'ichi Takeda was studying at IP. I believe that this introduction
was passed on to the professors at the National Institute of
Neuroscience NCNP, including Dr. Yo-ichi Nabeshima, who was Margaret's
colleague, and led to my return to Japan.

Finally, it is with great regret that I lost Professor Fran\c{c}ois
Gros on February 18, 2022. I thank him very much for receiving me in
his laboratory in IP and supporting my stay in France (his letter of
recommendation was required in order to obtain a residence permission
in France), and for his letter of recommendation to the National
Institute of Neuroscience NCNP, when I returned home. I am also
grateful to him for giving me the opportunity to hold the workshop
between France and Japan, and having the opportunity to speak at his
90th birthday celebration held at IP in April 2015. On April 24, 2023,
a farewell meeting and celebration in his memory was held at IP and the
French Academy of Sciences, which I was able to attend, after which I
would like to once again offer my sincere prayers for his soul.

\section*{Declaration of interests}
The authors do not work for, advise, own shares in, or receive funds
from any organization that could benefit from this article, and have
declared no affiliations other than their research organizations.

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