.).

Whole-mount immunofluorescence
Immunofluorescence was performed as previously described [20]. Briefly, zebrafish embryos were fixed with 4% PFA at 4°C overnight, washed 10 times with PBS for 5 min each wash, incubated with acetone at − 20°C for 10 min, and finally washed 10 times with PBS for 5 min each wash. Samples were blocked with 5% BSA containing 0.1% triton X-100, and incubated with phalloidin (Sigma, P5282, 1;2000) or Huc (Abcam, ab210554, 1: 250) at 4°C overnight. The embryos were then washed, and DAPI was used for nucleic acid staining. The embryos were examined by a Zeiss confocal laser-scanning microscope.
Additional file 1: Figure S1. V111G and G214S mutations alter subcellular localization of GMPPB. C2C12 myoblasts were transfected with HA-GMPPB WT, HA-GMPPB V111G, HA-GMPPB G214S or HA-GMPPB D334N. Compared to WT GMPPB, V111G mutant exhibited increased localization in the nucleus, while G214S mutant seemed to lose nuclear localization. D334N, previously reported to form aggregates within the cytoplasm, was used as a control. Scale bar: 10 μm. Figure S2. GMPPB KD causes muscle defects in zebra fish. (a) Phalloidin staining (red) of filamentous actin in zebra fish injected with control MO (NC), or gmppb MO at one-cell stage. Scale bar: 250 μm (b) Phalloidin staining (green) on the cross-sections of 24 hpf embryos injected with control MO (NC), or gmppb MO at one-cell stage. Scale bar: 100 μm. Figure S3. GMPPB V111G mutant fails to rescue muscle defects in zebra fish caused by GMPPB KD. Phalloidin (green) and DAPI (blue) staining on the crosssections of 48 hpf embryos injected with control MO (NC), or gmppb MO alone or together with mRNA encoding GMPPB WT or its mutants at one-cell stage. Scale bar: 100 μm.Russia is winning, despite billions of dollars in American and NATO military support to Ukraine, the Russians have already achieved their most important strategic objectives. Russian President Vladimir Putin did not simply wake up one day and decide to send troops into Ukrainian cities, he publicly warned the West about this plan of action years in advance, yet he was ignored by most western leaders. Russia did not come into Ukraine to lose, it is undoubtably obvious that after 66 days of warfare, Russia is defeating not only the Ukrainians but NATO and the United States as well, it is quite an achievement.
There is no way Russia is going to “lose” the battle in Ukraine, those who know the Russian military are quite aware that Putin has not used the full Russian might, in the eyes of the Russian President, they have not even begun to fight a real war. The Russians describe the conflict as a special military operation, the first phase of the Russian operation employed young officers undergoing new training methods, it was a bit of a test, despite heavy loses the Russian objective succeeded in the first month. Putin publicly addressed these young soldiers in a video a week into the conflict, he said “I know you are learning new complex methods of warfare”, despite the learning curve, the Russians have achieved their goals in much of the South and East of Ukraine.
The Russian federation has made it clear they are conducting a limited special military operation, a very specific plan to demilitarize and de-Nazi-fy Ukrainian cities. Russia still has not defined their operation as a “war” yet and if they do decide to declare war, we can expect an assault on Ukraine beyond our imagination. The Western mainstream media has sold us complete fabrications, nothing about this conflict has been based in journalistic truth, the narrative that Ukraine is “winning” is simply not accurate. We have been told Vladimir Putin is depressed, angry, frustrated, physically ill, demented, cruel, ordering the fatality of civilians, yet not one of these accusations have been based in any verifiable evidence. The Russian President has methodically taken control of the major trade routes, essentially choking the life out of Ukraine, he is winning. Putin took little port city Mariupol while the whole world had their eyes on the capital Kyiv, it was a brilliant move. Vladimir Putin has not revealed on single thing about his military operation in terms of strategy, the West still cannot figure him out.
Everything we learn about Russia comes from former generals on American TV, MSNBC, and CNN, they are almost always wrong. Russian Foreign minister Sergi Lavrov mocked the Americans trying to analyze the Russian military operation, he said “They don’t understand us, they don’t understand the Russian mindset, they don’t understand how we work”, in essence, Lavrov was making it clear everything the Western media prints about the war is a fabrication. The average Ukrainian citizen understands Putin better than any American, British, or German news analyst, in this aspect Russia is defeating the Americans, NATO and the Ukrainians.
Who is really fighting against Russia?
So far, the Russians have documented at least 40 different countries who are engaged in battle against them, even the Ukrainians bragged about how many hired contractors are battling the Russians. In the week since President of Ukraine Zelensky announced the creation of an ‘International Legion’ to defend Ukraine, 20,000 people from 52 countries have volunteered to fight in Ukraine, the country’s, Foreign Minister, Dmytro Kuleba said on television. On the Russian side are simply young Russians boys and Chechen (Pro Russian nationals) soldiers, that’s it. A simple look at a map will show how Russia has taken the key areas of Ukraine. The red and light red shows how Ukraine has already lost the battle.
Russia is not only winning against the Ukrainians, but they are also defeating NATO and American money/weapons, the U.S loves funding foreign wars but they don’t know how to win them.
Britain, Germany. Italy and France want no part of war with Russia, Western Europe would never send a dollar (or any military equipment) to Ukraine without the United States coercing them to do so. Western Europeans are not in the least bit interested in engaging in war in Eastern Europe, most Westerners are worried about unemployment, food shortage and inflation, not Ukraine.
Our media is not allowed to tell you Russia is winning.
According to our biased media, cities like Mariupol and Kherson are not under Russian control, they tell the public there is “fierce resistance” to Russian attacks, this is of course a lie. A month ago, a lifelong resident of Kherson reported that the city was in 100% Russian control, but public is not allowed to know this truth.
See video: This would never air on American television.
The guy on camera tells the truth “Kherson is under 100% Russian control”, the reporter did not expect that answer she tries to cover the truth, she says “there are conflicting reports on who controls the city”, then he dropped the full truth again. Russia is in 100% control; this was a rare truth that slipped by Pro-Ukraine media.
Finding honest news is almost impossible today, the fact of the matter is that Russia is not only winning, but they are also winning relatively easily despite suffering heavy losses. Does anyone really believe that Russia could not take all of Ukraine if they want it? At the start of the conflict every American media pundit said Russia would take the capital city Kyiv in less than a week, yet Russia “chose” not to do so. No one can figure out Putin’s war strategy. The Ukrainian civilians are sadly being slaughtered, they are caught between the Russian military might and Zelensky’s ego. No matter how many thousands of civilians slaughtered, Washington has only one response…more weapons, more weapons, more weapons.
The United States Government does not care the least bit about Ukrainian civilians, Ukraine (which is actually a welfare state) has never stood on its own two feet; it is a beggar state. The President of Ukraine is not intelligent, but he is slick, he knows how to milk every dollar out of the United States. It is a cruel game; Washington relies on the American mainstream media to keep showing bombed out Ukrainian children on the news to justify the billions of dollars we keep sending to Zelensky. Ukraine is one of the most corrupt states in the world, dumping billions of dollars worth of weapons into the land is lunacy, yet we will keep on doing it. The bottom line is that Russia is winning, and nothing will change this outcome, no amount of weapons dump into Ukraine is going to change the result of the war. No one in Washington cares about the Ukrainian civilians, the United States doesn’t care, Zelensky doesn’t care and certainly NATO does not care; if they did care they would have told Ukrainians to lay down arms 2 months ago and tried to reason (negotiate) with Putin.
We are defending Ukraine as if it is one of our 50 states, we are taunting a nuclear superpower, we are encouraging the hotheaded Zelensky, all while our corrupt American media leads the drumbeats to world war 3.
Too many Americans do not know their press is controlled for propaganda purposes.
There is a good case to be made that the NATO trained Ukraine forces in the southeast are the very best that NATO can muster. They have been trained for 8 years. Some fought in 2014. Many have been operating that equipment for years shelling the