5.1  Conclusion.  


During   the   last   decades,   a   vast   amount   of   publications   have   unraveled   the   importance  of  mitochondrial  dynamics  during  cells’s  life  and  death.  Nowadays,  it   is   clear   that   the   control   of   mitochondrial   shape   through   the   balance   of   the   opposite   processes   of   fusion   and   fission   is   essential   in   mitochondrial   functions   such   as   respiration   and   cell   death.   However,   many   aspects   of   the   mechanisms   underlying  mitochondrial  dynamics  are  yet  unclear.  In  particular,  mitochondrial   fission  was  though  to  be  a  two-­‐component  system,  with  Fis1  being  the  receptor   for   the   translocation   of   Drp1   to   the   OMM.   The   discovery   of   Mff   and   MiDs   as   receptors   for   Drp1   in   mammals   challenged   the   relevance   of   Fis1   in   mitochondrial  fission.  In  this  project,  we  found  compelling  evidence  supporting   the  importance  of  Fis1  gene  in  mitochondrial  morphology  with  consequences  in   mitochondrial   functionality   and   cell   death.   Additionally,   we   report   the   unexpected   discovery   that   Fis1   is   alternatively   spliced   in   three   variants   expressed   in   several   tissues.   While   two   of   these   variants   play   a   role   in   mitochondrial  fission,  increased  mFis1.2  levels  cause  mitochondrial  elongation.  

By  mapping  the  relative  expression  of  mFis1.2  in  pathophysiological  conditions,   we   found   it   increased   during   macroautophagy   in   a   PKA-­‐dependent   manner   to   produce   mitochondrial   elongation.   Hence,   we   have   identifying   a   novel   mechanism   for   mitochondrial   morphology   changes   during   autophagy   that,  

together   with   the   already   known   mechanism   of   PKA-­‐dependent   inhibition   of   Drp1,  helps  to  enhance  respiration  in  conditions  of  nutrient  deprivation.  


5.2  Perspectives.  


Great  emphasis  has  been  placed  in  the  many  post-­‐translational  modification  that   Drp1   suffers   (phosphorylation,   ubiquitination   and   sumoylation)   and   how   they   modulate   mitochondrial   fission.   On   the   other   hand,   Drp1   receptors   have   been   treated   as   invariable   proteins,   ready   to   interact   with   Drp1   whenever   it   is   activated.   The   results   presented   in   this   work   highlight   the   relevance   of   the   modulation  of  Fis1  variant  expression,  especially  of  mFis1.2,  in  the  regulation  of   mitochondrial  morphology.  Likewise,  other  fission  factors  may  be  regulated  in  a   similar  manner.  For  example,  Mff  also  has  splice  variants  that  may  have  different   expression  according  to  internal  or  external  cues  as  we  showed  for  mouse  Fis1.  

It   is   interesting   to   note   that   only   a   small   deletion   in   the   amino-­‐terminus   of   mFsi1.2   induces   an   apposite   effect   on   mitochondrial   morphology,   given   to   the   coiled   coil   domain   in   this   region   a   regulatory   role   in   mitochondrial   dynamics.  

However,  whether  the  coiled  coil  arm  is  important  in  the  interaction  of  Fis1  with   itself   or   with   other   mitochondrial   shaping   proteins   still   has   to   be   investigated.  

Additionally,  human  Fis1  gene  is  also  able  to  produce  four  splice  variants,  some   with   differences   in   the   coiled   coil   arm,   however,   nothing   is   known   about   their   expression   pattern   and   regulation   nor   about   their   role   in   mitochondrial   morphology.  Hence,  it  is  possible  that  mitochondrial  shape  in  human  cells  may   also   be   regulated   by   Fis1   splice   variants.   To   this   regard,   it   is   of   paramount   importance   to   direct   studies   in   mouse   and   human   cells   aimed   to   address   the  

  121   hierarchical   interaction   between   the   different   Drp1   receptors   and   their   splice   variants   in   order   to   produce   a   specific   mitochondrial   shape   in   response   to   a   given   stimulus.   Most   likely,   a   detailed   mapping   of   the   mitochondrial   fission   transcriptome   and   interactome   will   contribute   to   a   clearer   view   of   how   mitochondrial  fragmentation  is  achieved.  

Finally,   mitochondria   have   a   central   role   in   cancer   and   neurodegenerative   diseases  and  data  in  this  work  clearly  show  that  mFis1  impinges  not  only  in  the   efficiency   of   mitochondrial   respiration   but   as   well   in   the   progression   of   cell   death.  Future  work  in  animal  models  should  clarify  whether  the  modulation  of   mFis1  variants  expression  could  have  an  impact  in  physiological  processes  with   pathological  consequences.  




Chapter  6  

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